Methods and compostions for treating proliferative diseases

ABSTRACT

The present invention provides combination therapy methods of treating proliferative diseases (such as cancer) comprising a first therapy comprising administering to an individual an effective amount of a taxane in a nanoparticle composition, and a second therapy which may include, for example, radiation, surgery, administration of chemotherapeutic agents, or combinations thereof. Also provided are methods of administering to an individual a drug taxane in a nanoparticle composition based on a metronomic dosing regime.

RELATED APPLICATIONS

This application is a divisional application of U.S. patent applicationSer. No. 11/544,242, filed on Oct. 6, 2006, which is a continuationapplication of U.S. patent application Ser. No. 11/359,286, filed onFeb. 21, 2006, which claims priority benefit to the provisionalapplication 60/654,245, filed on Feb. 18, 2005, the content of each ofwhich are incorporated by reference herein in their entirety.

TECHNICAL FIELD

The present invention relates to methods and compositions for thetreatment of proliferative diseases comprising the administration of acombination of a taxane and at least one other and other therapeuticagents, as well as other treatment modalities useful in the treatment ofproliferative diseases. In particular, the invention relates to the useof nanoparticles comprising paclitaxel and albumin (such as Abraxane™)in combination with other chemotherapeutic agents or radiation, whichmay be used for the treatment of cancer.

BACKGROUND

The failure of a significant number of tumors to respond to drug and/orradiation therapy is a serious problem in the treatment of cancer. Infact, this is one of the main reasons why many of the most prevalentforms of human cancer still resist effective chemotherapeuticintervention, despite certain advances in the field of chemotherapy.

Cancer is now primarily treated with one or a combination of three typesof therapies: surgery, radiation, and chemotherapy. Surgery is atraditional approach in which all or part of a tumor is removed from thebody. Surgery generally is only effective for treating the earlierstages of cancer. While surgery is sometimes effective in removingtumors located at certain sites, for example, in the breast, colon, andskin, it cannot be used in the treatment of tumors located in otherareas, inaccessible to surgeons, nor in the treatment of disseminatedneoplastic conditions such as leukemia. For more than 50% of cancerindividuals, by the time they are diagnosed they are no longercandidates for effective surgical treatment. Surgical procedures mayincrease tumor metastases through blood circulation during surgery. Mostof cancer individuals do not die from the cancer at the time ofdiagnosis or surgery, but rather die from the metastasis and therecurrence of the cancer.

Other therapies are also often ineffective. Radiation therapy is onlyeffective for individuals who present with clinically localized diseaseat early and middle stages of cancer, and is not effective for the latestages of cancer with metastasis. Radiation is generally applied to adefined area of the subject's body which contains abnormal proliferativetissue, in order to maximize the dose absorbed by the abnormal tissueand minimize the dose absorbed by the nearby normal tissue. However, itis difficult (if not impossible) to selectively administer therapeuticradiation to the abnormal tissue. Thus, normal tissue proximate to theabnormal tissue is also exposed to potentially damaging doses ofradiation throughout the course of treatment. There are also sometreatments that require exposure of the subject's entire body to theradiation, in a procedure called “total body irradiation”, or “TBI.” Theefficacy of radiotherapeutic techniques in destroying abnormalproliferative cells is therefore balanced by associated cytotoxiceffects on nearby normal cells. Because of this, radiotherapy techniqueshave an inherently narrow therapeutic index which results in theinadequate treatment of most tumors. Even the best radiotherapeutictechniques may result in incomplete tumor reduction, tumor recurrence,increasing tumor burden, and induction of radiation resistant tumors.

Chemotherapy involves the disruption of cell replication or cellmetabolism. Chemotherapy can be effective, but there are severe sideeffects, e.g., vomiting, low white blood cells (WBC), loss of hair, lossof weight and other toxic effects. Because of the extremely toxic sideeffects, many cancer individuals cannot successfully finish a completechemotherapy regime. Chemotherapy-induced side effects significantlyimpact the quality of life of the individual and may dramaticallyinfluence individual compliance with treatment. Additionally, adverseside effects associated with chemotherapeutic agents are generally themajor dose-limiting toxicity (DLT) in the administration of these drugs.For example, mucositis is one of the major dose limiting toxicity forseveral anticancer agents, including the antimetabolite cytotoxic agents5-FU, methotrexate, and antitumor antibiotics, such as doxorubicin. Manyof these chemotherapy-induced side effects if severe may lead tohospitalization, or require treatment with analgesics for the treatmentof pain. Some cancer individuals die from the chemotherapy due to poortolerance to the chemotherapy. The extreme side effects of anticancerdrugs are caused by the poor target specificity of such drugs. The drugscirculate through most normal organs of individuals as well as intendedtarget tumors. The poor target specificity that causes side effects alsodecreases the efficacy of chemotherapy because only a fraction of thedrugs is correctly targeted. The efficacy of chemotherapy is furtherdecreased by poor retention of the anti-cancer drugs within the targettumors.

Due to the severity and breadth of neoplasm, tumor and cancer, there isa great need for effective treatments of such diseases or disorders thatovercome the shortcomings of surgery, chemotherapy, and radiationtreatment.

Problems of Chemotherapeutic Agents

The drug resistance problem is a reason for the added importance ofcombination chemotherapy, as the therapy both has to avoid the emergenceof resistant cells and to kill pre-existing cells which are already drugresistant.

Drug resistance is the name given to the circumstance when a diseasedoes not respond to a treatment drug or drugs. Drug resistance can beeither intrinsic, which means the disease has never been responsive tothe drug or drugs, or it can be acquired, which means the disease ceasesresponding to a drug or drugs that the disease had previously beenresponsive to. Multidrug resistance (MDR) is a specific type of drugresistance that is characterized by cross-resistance of a disease tomore than one functionally and/or structurally unrelated drugs.Multidrug resistance in the field of cancer is discussed in greaterdetail in “Detoxification Mechanisms and Tumor Cell Resistance toAnticancer Drugs,” by Kuzmich and Tew, particularly section VII “TheMultidrug-Resistant Phenotype (MDR),” Medical Research Reviews, Vol. 11,No. 2, 185-217, (Section VII is at pp. 208-213) (1991); and in“Multidrug Resistance and Chemosensitization: Therapeutic Implicationsfor Cancer Chemotherapy,” by Georges, Sharom and Ling, Advances inPharmacology, Vol. 21, 185-220 (1990).

One form of multi-drug resistance (MDR) is mediated by a membrane bound170-180 kD energy-dependent efflux pump designated as P-glycoprotein(P-gp). P-glycoprotein has been shown to play a major role in theintrinsic and acquired resistance of a number of human tumors againsthydrophobic, natural product drugs. Drugs that act as substrates for andare consequently detoxified by P-gp include the vinca alkaloids(vincristine and vinblastine), anthracyclines (Adriamycin), andepipodophyllotoxins (etoposide). While P-gp associated MDR is a majordeterminant in tumor cell resistance to chemotherapeutic agents, it isclear that the phenomenon of MDR is multifactorial and involves a numberof different mechanisms.

A major complication of cancer chemotherapy and of antiviralchemotherapy is damage to bone marrow cells or suppression of theirfunction. Specifically, chemotherapy damages or destroys hematopoieticprecursor cells, primarily found in the bone marrow and spleen,impairing the production of new blood cells (granulocytes, lymphocytes,erythrocytes, monocytes, platelets, etc.). Treatment of cancerindividuals with 5-fluorouracil, for example, reduces the number ofleukocytes (lymphocytes and/or granulocytes), and can result in enhancedsusceptibility of the. individuals to infection. Many cancer individualsdie of infection or other consequences of hematopoietic failuresubsequent to chemotherapy. Chemotherapeutic agents can also result insubnormal formation of platelets which produces a propensity towardhemorrhage. Inhibition of erythrocyte production can result in anemia.For some cancer individuals, the risk of damage to the hematopoieticsystem or other important tissues frequently limits the opportunity forchemotherapy dose escalation of chemotherapy agents high enough toprovide good antitumor or antiviral efficacy. Repeated or high dosecycles of chemotherapy may be responsible for severe stem cell depletionleading to serious long-term hematopoietic sequelea and marrowexhaustion.

Prevention of, or protection from, the side effects of chemotherapywould be a great benefit to cancer individuals. For life-threateningside effects, efforts have concentrated on altering the dose andschedules of the chemotherapeutic agent to reduce the side effects.Other options are becoming available, such as the use of granulocytecolony stimulating factor (G-CSF), granulocyte-macrophage-CSF (GM-CSF),epidermal growth factor (EGF), interleukin 11, erythropoietin,thrombopoietin, megakaryocyte development and growth factor, pixykines,stem cell factor, FLT-ligand, as well as interleukins 1, 3, 6, and 7, toincrease the number of normal cells in various tissues before the startof chemotherapy (See Jimenez and Yunis, Cancer Research 52:413-415;1992). The mechanisms of protection by these factors, while not fullyunderstood, are most likely associated with an increase in the number ofnormal critical target cells before treatment with cytotoxic agents, andnot with increased survival of cells following chemotherapy.

Chemotherapeutic Targeting For Tumor Treatment

Both the growth and metastasis of solid tumors areangiogenesis-dependent (Folkman, J. Cancer Res., 46, 467-73 (1986);Folkman, J. Nat. Cancer Inst., 82, 4-6 (1989); Folkman et al., “TumorAngiogenesis,” Chapter 10, pp. 206-32, in The Molecular Basis of Cancer,Mendelsohn et al., eds. (W. B. Saunders, 1995)). It has been shown, forexample, that tumors which enlarge to greater than 2 mm in diameter mustobtain their own blood supply and do so by inducing the growth of newcapillary blood vessels. After these new blood vessels become embeddedin the tumor, they provide nutrients and growth factors essential fortumor growth as well as a means for tumor cells to enter the circulationand metastasize to distant sites, such as liver, lung or bone (Weidner,New Eng. J. Med., 324(1), 1-8 (1991)). When used as drugs intumor-bearing animals, natural inhibitors of angiogenesis can preventthe growth of small tumors (O'Reilly et al., O'Reilly et al., Cell, 79,315-28 (1994)). Indeed, in some protocols, the application of suchinhibitors leads to tumor regression and dormancy even after cessationof treatment (O'Reilly et al., Cell, 88, 277-85 (1997)). Moreover,supplying inhibitors of angiogenesis to certain tumors can potentiatetheir response to other therapeutic regimes (e.g., chemotherapy) (see,e.g., Teischer et al., Int. J. Cancer, 57, 920-25 (1994)).

Protein tyrosine kinases catalyze the phosphorylation of specifictyrosyl residues in various proteins involved in the regulation of cellgrowth and differentiation (A. F. Wilks, Progress in Growth FactorResearch, 1990, 2, 97-111; S. A. Courtneidge, Dev. Supp. 1, 1993, 57-64;J. A. Cooper, Semin. Cell Biol., 1994, 5(6), 377-387; R. F. Paulson,Semin. Immunol., 1995, 7(4), 267-277; A. C. Chan, Curr. Opin. Immunol.,1996, 8(3), 394-401). Protein tyrosine kinases can be broadly classifiedas receptor (e.g. EGFr, c-erbB-2, c-met, tie-2, PDGFr, FGFr) ornon-receptor (e.g. c-src, Ick, Zap70) kinases. Inappropriate oruncontrolled activation of many of these kinases, i.e. aberrant proteintyrosine kinase activity, for example by over-expression or mutation,has been shown to result in uncontrolled cell growth. For example,elevated epidermal growth factor receptor (EGFR) activity has beenimplicated in non-small cell lung, bladder and head and neck cancers,and increased c-erbB-2 activity in breast, ovarian, gastric andpancreatic cancers. Thus, inhibition of protein tyrosine kinases shouldbe useful as a treatment for tumors such as those outlined above.

Growth factors are substances that induce cell proliferation, typicallyby binding to specific receptors on cell surfaces. Epidermal growthfactor (EGF) induces proliferation of a variety of cells in vivo, and isrequired for the growth of most cultured cells. The EGF receptor is a170-180 kD membrane-spanning glycoprotein, which is detectable on a widevariety of cell types. The extracellular N-terminal domain of thereceptor is highly glycosylated and binds EGF antibodies thatselectively bind to EGFR. Agents that competitively bind to EGFR havebeen used to treat certain types of cancer, since many tumors ofmesodermal and ectodermal origin overexpress the EGF receptor. Forexample, the EGF receptor has been shown to be overexpressed in manygliomas, squamous cell carcinomas, breast carcinomas, melanomas,invasive bladder carcinomas and esophageal cancers. Attempts to exploitthe EGFR system for anti-tumor therapy have generally involved the useof monoclonal antibodies against the EGFR. In addition, studies withprimary human mammary tumors have shown a correlation between high EGFRexpression and the presence of metastases, higher rates ofproliferation, and shorter individual survival.

Herlyn et al., in U.S. Pat. No. 5,470,571, disclose the use ofradiolabeled Mab 425 for treating gliomas that express EGFR. Herlyn etal. report that anti-EGFR antibodies may either stimulate or inhibitcancer cell growth and proliferation. Other monoclonal antibodies havingspecificity for EGFR, either alone or conjugated to a cytotoxiccompound, have been reported as being effective for treating certaintypes of cancer. Bendig et al, in U.S. Pat. No. 5,558,864, disclosetherapeutic anti-EGFR Mab's for competitively binding to EGFR. Heimbrooket al., in U.S. Pat. No. 5,690,928, disclose the use of EGF fused to aPseudomonas species-derived-endotoxin for the treatment of bladdercancer. Brown et al., in U.S. Pat. No. 5,859,018, disclose a method fortreating diseases characterized by cellular hyperproliferation mediatedby, inter alia, EGF.

Chemotherapeutic Modes of Administration

People diagnosed as having cancer are frequently treated with single ormultiple chemotherapeutic agents to kill cancer cells at the primarytumor site or at distant sites to where cancer has metastasized.Chemotherapy treatment is typically given either in a single or inseveral large doses or over variable times of weeks to months. However,repeated or high dose cycles of chemotherapy may be responsible forincreased toxicities and severe side effects.

New studies suggest that metronomic chemotherapy, the low-dose andfrequent administration of cytotoxic agents without prolonged drug-freebreaks, targets activated endothelial cells in the tumor vasculature. Anumber of preclinical studies have demonstrated superior anti-tumorefficacy, potent antiangiogenic effects, and reduced toxicity and sideeffects (e.g., myelosuppression) of metronomic regimes compared tomaximum tolerated dose (MTD) counterparts (Bocci, et al., Cancer Res,62:6938-6943, (2002); Bocci, et al., PNAS, vol. 100(22):12917-12922,(2003); and Bertolini, et al., Cancer Res, 63(15):4342-4346, (2003)). Itremains unclear whether all chemotherapeutic drugs exert similar effectsor whether some are better suited for such regimes than others.Nevertheless, metronomic chemotherapy appears to be effective inovercoming some of the major shortcomings associated with chemotherapy.

Chemotherapeutic Agents

Paclitaxel has been shown to have significant antineoplastic andanticancer effects in drug-refractory ovarian cancer and has shownexcellent antitumor activity in a wide variety of tumor models, and alsoinhibits angiogenesis when used at very low doses (Grant et al., Int. J.Cancer, 2003). The poor aqueous solubility of paclitaxel, however,presents a problem for human administration. Indeed, the delivery ofdrugs that are inherently insoluble or poorly soluble in an aqueousmedium can be seriously impaired if oral delivery is not effective.Accordingly, currently used paclitaxel formulations (e.g., Taxol®)require a Cremophor® to solubilize the drug. The presence of Cremophor®in this formulation has been linked to severe hypersensitivity reactionsin animals (Lorenz et al., Agents Actions 7:63-67 (1987)) and humans(Weiss et al., J. Clin. Oncol. 8: 1263-68 (1990)) and consequentlyrequires premedication of individuals with corticosteroids(dexamethasone) and antihistamines. It was also reported that clinicallyrelevant concentrations of the formulation vehicle Cremophor® EL inTaxol® nullify the antiangiogenic activity of paclitaxel, suggestingthat this agent or other anticancer drugs formulated in Cremophor® ELmay need to be used at much higher doses than anticipated to achieveeffective metronomic chemotherapy (Ng et al., Cancer Res., 64:821-824(2004)). As such, the advantage of the lack of undesirable side effectsassociated with low-dose paclitaxel regimes vs. conventional MTDchemotherapy may be compromised. See also U.S. Patent Pub. No.2004/0143004; WO00/64437.

Abraxane™ is a Cremophor® EL-Free Nanoparticle Albumin-bound Paclitaxel

Preclinical models have shown significant improvement in the safety andefficacy of Abraxane™ compared with Taxol® (Desai et al.,EORTC-NCI-AACR, 2004) and in individuals with metastatic breast cancer(O'Shaughnessy et al., San Antonio Breast Cancer Symposium, Abstract#1122, Dec. 2003). This is possibly due to the absence of surfactants(e.g., Cremophor® or Tween® 80, used in Taxol® and Taxotere®,respectively) in Abraxane™, and/or preferential utilization of analbumin-based transport mechanism utilizing gp60/caveolae onmicrovascular endothelial cells (Desai et al., EORTC-NCI-AACR, 2004). Inaddition, both Cremophor® and Tween® 80 have been shown to stronglyinhibit the binding of paclitaxel to albumin, possibly affecting albuminbased transport (Desai et al., EORTC-NCI-AACR, 2004).

IDN5109 (Ortataxel) is a new taxane, currently in phase II, selected forits lack of cross-resistance in tumor cell lines expressing themultidrug resistant phenotype (MDR/Pgp) and inhibition of P-glycoprotein(Pgp) (Minderman; Cancer Chemother. Pharmacol. 2004; 53:363-9). Due toits hydrophobicity, IDN5109 is currently formulated in the surfactantTween® 80 (same vehicle as Taxotere®). Removal of surfactants fromtaxane formulations e.g., in the case of nanoparticle albumin-boundpaclitaxel (Abraxane™) showed improvements in safety and efficacy overtheir surfactant containing counterparts (O'Shaughnessy et al., SanAntonio Breast Cancer Symposium, Abstract #1122, December 2003). Tween®80 also strongly inhibited the binding of the taxane, paclitaxel, toalbumin, possibly compromising albumin based drug transport via the gp60receptor on microvessel endothelial cells (Desai et al., EORTC-NCI-AACR,2004).

The antitumor activity of colchicine, which is the major alkaloid of theautumn crocus, Colchicum autumnale, and the African climbing lily,Gloriosa superba, was first reported at the beginning of the 20^(th)century. The elucidation of its structure was finally completed fromX-ray studies and a number of total syntheses (see Shiau et al., J.Pharm. Sci. 1978, 67(3) 394-397). Colchicine is thought to be a mitoticpoison, particularly in tyhmic, intestinal, and hermatopoietic cells,which acts as a spindle poison and blocks the kinesis. Its effect on themitotic spindle is thought to represent a special case of its effects onvarious organized, labile, fibrillar systems concerned with structureand movement.

Thiocolchicine dimer IDN5404 was selected for its activity in humanovarian subline resistant to cisplatin and topotecan A2780-CIS andA2780-TOP. This effect was related to dual mechanisms of action, i.e.,microtubule activity as in Vinca alkaloids and a topoisomerase Iinhibitory effect different from camptothecin. (Raspaglio, BiochemicalPharmacology 69:113-121 (2005)).

It has been found that nanoparticle compositions of a taxane (such asalbumin bound paclitaxel (Abraxane™)) have significantly lowertoxicities than other taxanes like Taxol® and Taxotere® withsignificantly improved outcomes in both safety and efficacy.

Combination chemotherapy, e.g., combining one or more chemotherapeuticagents or other modes of treatment, e.g., combining for example,chemotherapy with radiation or surgery and chemotherapy, have been foundto be more successful than single agent chemotherapeutics or individualmodes of treatment respectively.

Other references include U.S. Pub. No. 2006/0013819; U.S. Pub. No.2006/0003931; WO05/117986; WO05/117978; and WO05/000900.

More effective treatments for proliferative diseases, especially cancer,are needed.

The disclosures of all publications, patents, patent applications andpublished patent applications referred to herein are hereby incorporatedherein by reference in their entirety.

BRIEF SUMMARY OF THE INVENTION

The present invention provides methods for the treatment ofproliferative diseases such as cancer. The invention providescombination therapy methods of treating proliferative diseases (such ascancer), comprising a) a first therapy comprising administering to anindividual an effective amount of a composition comprising nanoparticlescomprising a taxane (such as paclitaxel) and a carrier protein (such asalbumin) and b) a second therapy, such as chemotherapy, radiationtherapy, surgery, or combinations thereof. In another aspect, there areprovided methods of administering to an individual a compositioncomprising nanoparticles comprising a taxane (such as paclitaxel) and acarrier protein (such as albumin) based on a metronomic dosing regime.

In some embodiments, the invention provides a method of treating aproliferative disease (such as cancer) in an individual comprisingadministering to the individual a) an effective amount of a compositioncomprising nanoparticles-comprising a taxane and a carrier protein (suchas albumin), and b) an effective amount of at least one otherchemotherapeutic agent. In some embodiments, the invention provides amethod of treating a proliferative disease (such as cancer) in anindividual comprising administering to the individual a) an effectiveamount of a composition comprising nanoparticies comprising paclitaxeland an albumin (such as Abraxane ™), and b) an effective amount of atleast one other chemotherapeutic agent. In some embodiments, thechemotherapeutic agent is any of (and in some embodiments selected fromthe group consisting of) antimetabolites (including nucleoside analogs),platinum-based agents, alkylating agents, tyrosine kinase inhibitors,anthracycline antibiotics, vinca alkloids, proteasome inhibitors,macrolides, and topoisomerase inhibitors. In some embodiments, thechemotherapeutic agent is a platinum-based agent, such as carboplatin.

In some embodiments, the composition comprising nanoparticles (alsoreferred to as “nanoparticle composition”) and the chemotherapeuticagent are administered simultaneously, either in the same composition orin separate compositions. In some embodiments, the nanoparticlecomposition and the chemotherapeutic agent are administeredsequentially, i.e., the nanoparticle composition is administered eitherprior to or after the administration of the chemotherapeutic agent. Insome embodiments, the administration of the nanoparticle composition andthe chemotherapeutic agent are concurrent, i.e., the administrationperiod of the nanoparticle composition and that of the chemotherapeuticagent overlap with each other. In some embodiments, the administrationof the nanoparticle composition and the chemotherapeutic agent arenon-concurrent. For example, in some embodiments, the administration ofthe nanoparticle composition is terminated before the chemotherapeuticagent is administered. In some embodiments, the administration of thechemotherapeutic agent is terminated before the nanoparticle compositionis administered.

In some embodiments, the first therapy taxane is nano-particle albuminbound paxlitaxel, described, for example, in U.S. Pat. No. 6,566,405,and commercially available under the tradename Abraxane™. In addition,the first therapy taxane is also considered to be nanoparticle albuminbound docetaxel-described for example in U.S. Patent ApplicationPublication 2005/0004002A1.

In another aspect, there is provided a method of treating aproliferative disease (such as cancer) in an individual comprising a) afirst therapy comprising administering to the individual a compositioncomprising nanoparticles comprising a taxane and a carrier protein (suchas albumin), and b) a second therapy comprising radiation therapy,surgery, or combinations thereof. In some embodiments, there is provideda method of treating a proliferative disease (such as cancer) in anindividual comprising a) a first therapy comprising administering to theindividual a composition comprising nanoparticles comprising paclitaxeland an albumin (such as Abraxane™), and b) a second therapy comprisingradiation therapy, surgery, or combinations thereof In some embodiments,the second therapy is radiation therapy. In some embodiments, the secondtherapy is surgery. In some embodiments, the first therapy is carriedout prior to the second therapy. In some embodiments, the first therapyis carried out after the second therapy.

In another aspect, the method comprises administering to a mammal havinga proliferative disease (such as cancer) a combination therapycomprising a first therapy comprising a taxane and a second therapyselected from the group consisting of chemotherapeutic agent andradiation or combinations thereof. The combination therapy may beadministered in any of a variety of ways such as sequentially orsimultaneously, and if sequential, the taxane may be administered beforeor after the second therapy although it is preferred that the firsttherapy comprising a taxane is administered first. It will also beunderstood that the second therapy can include more than one.chemotherapeutic agent.

The present invention also provides metronomic therapy regimes. In someembodiments, there is provided a method of administering a compositioncomprising nanoparticles comprising a taxane and a carrier protein (suchas albumin), wherein the nanoparticle composition is administered over aperiod of at least one month, wherein the interval between eachadministration is no more than about a week, and wherein the dose oftaxane at each administration is about 0.25% to about 25% of its maximumtolerated dose following a traditional dosing regime. In someembodiments, there is provided a method of administering a compositioncomprising nanoparticles comprising paclitaxel and an albumin (such asAbraxane™), wherein the nanoparticle composition is administered over aperiod of at least one month, wherein the interval between eachadministration is no more than about a week, and wherein the dose ofpaclitaxel at each administration is about 0.25% to about 25% of itsmaximum tolerated dose following a traditional dosing regime. In someembodiments, the dose of the taxane (such as paclitaxel, for exampleAbraxane™) per administration is less than about any of 1%, 2%, 3%, 4%,5%, 6%, 7%, 8%, 9%, 10%, 11%, 12%, 13%, 14%, 15%, 18%, 20%, 22%, 24%, or25% of the maximum tolerated dose. In some embodiments, the nanoparticlecomposition is administered at least about any of 1×, 2×, 3×, 4×, 5×,6×, 7× (i.e., daily) a week. In some embodiments, the intervals betweeneach administration are less than about any of 7 days, 6 days, 5 days, 4days, 3 days, 2 days, and 1 day. In some embodiments, the nanoparticlecomposition is administered over a period of at least about any of 2, 3,4, 5, 6, 7, 8, 9, 10, 11, 12, 18, 24, 30 and 36 months.

In some embodiments, there is provided a method of administering acomposition comprising nanoparticles comprising a taxane and a carrierprotein (such as albumin), wherein the taxane is administered over aperiod of at least one month, wherein the interval between eachadministration is no more than about a week, and wherein the dose of thetaxane at each administration is about 0.25 mg/m² to about 25 mg/m². Insome embodiments, there is provided a method of administering acomposition comprising nanoparticles comprising paclitaxel and analbumin (such as Abraxane™) and a carrier protein (such as albumin),wherein the paclitaxel is administered over a period of at least onemonth, wherein the interval between each administration is no more thanabout a week, and wherein the dose of the taxane at each administrationis about 0.25 mg/m² to about 25 mg/m². In some embodiments, the dose ofthe taxane (such as paclitaxel, for example Abraxane™) peradministration is less than about any of 2, 3, 4, 5, 6, 7, 8, 9, 10, 11,12, 13, 14, 15, 18, 20, 22, and 25 mg/m². In some embodiments, thenanoparticle composition is administered at least about any of 1×, 2×,3×, 4×, 5×, 6×, 7× (i.e., daily) a week. In some embodiments, theintervals between each administration are less than about any of 7 days,6 days, 5 days, 4 days, 3 days, 2 days, and 1 day. In some embodiments,the nanoparticle composition is administered over a period of at leastabout any of 2, 3, 4, 5, 6, 7, 8, 9, 10, 11, 12, 18, 24, 30 and 36months.

The methods of the invention generally comprise administration of acomposition comprising nanoparticles comprising a taxane and a carrierprotein. In some embodiments, the nanoparticle composition comprisesnanoparticles comprising paclitaxel and an albumin. In some embodiments,the paclitaxel/albumin nanoparticles have an average diameter of nogreater than about 200 nm. In some embodiments, the paclitaxel/albuminnanoparticle composition is substantially free (such as free) ofsurfactant (such as Cremophor). In some embodiments, the weight ratio ofthe albumin to paclitaxel in the composition is about 18:1 or less, suchas about 9:1 or less. In some embodiments, the paclitaxel is coated withalbumin. In some embodiments, the paclitaxel/albumin nanoparticles havean average diameter of no greater than about 200 nm and thepaclitaxel/albumin composition is substantially free (such as free) ofsurfactant (such as Cremophor). In some embodiments, thepaclitaxel/albumin nanoparticles have an average diameter of no greaterthan about 200 nm and the paclitaxel is coated with albumin. Othercombinations of the above characteristics are also contemplated. In someembodiments, the nanoparticle composition is Abraxane™. Nanoparticlecompositions comprising other taxanes (such as docetaxel and ortataxel)may also comprise one or more of the above characteristics.

These and other aspects and advantages of the present invention willbecome apparent from the subsequent detailed description and theappended claims. It is to be understood that one, some, or all of theproperties of the various embodiments described herein may be combinedto form other embodiments of the present invention.

BRIEF DESCRIPTION OF FIGURES

FIG. 1A shows the effect of ABI-007 on rat aortic ring angiogenesis.FIG. 1B shows the effect of ABI-007 on human endothelial cellproliferation. FIG. 1C shows the effect of ABI-007 on endothelial celltube formation.

FIG. 2 shows the determination of an optimal biological dose of ABI-007for metronomic dosing. Shown are the levels of viable circulatingendothelial progenitors (CEPs) in peripheral blood of Balb/cJ mice inresponse to escalating doses of ABI-007. Untr'd, untreated control; S/A,saline/albumin vehicle control. Bars, mean±SE. * Significantly (p<0.05 )different from the untreated control.

FIGS. 3A and 3B show the effects of ABI-007 and Taxol used in metronomicor MTD regimes on MDA-MB-23 1 (A) and PC3 (B) tumor growth tumor-bearingSCID mice. FIGS. 3C and 3D show the effects of ABI-007 and Taxol used inmetronomic or MTD regimes on the body weight of MDA-MB-23 1 (C) and PC3(D) tumor-bearing SCID mice.

FIGS. 4A and 4B show changes in the levels of viable circulatingendothelial progenitors (CEPs) in peripheral blood of MDA-MB-23 1 (FIG.4A) and PC3 (FIG. 4B) tumor-bearing SCID mice after treatment with A,saline/albumin; B, Cremophor EL control; C, metronomic Taxol 1.3 mg/kg;D, E, and F, metronomic ABI-007 3, 6, and 10 mg/kg, respectively; G, MTDTaxol; H, MTD ABI-007. Bars, mean±SE. ^(a) Significantly (p<0.05)different from saline/albumin vehicle control. ^(b) Significantly(p<0.05) different from Cremophor EL vehicle control.

FIG. 5A shows intratumoral microvessel density of MDA-MB-231 (▪) and PC3(□) xenografts treated with A, saline/albumin; B, Cremophor EL control;C, metronomic Taxol 1.3 mg/kg; D, E, and F, metronomic ABI-007 3, 6, and10 mg/kg, respectively; G, MTD Taxol; H, MTD ABI-007. Bars, mean +SE.FIG. 5B and 5C show the correlation between intratumoral microvesseldensity and the number of viable CEPs in peripheral blood in MDA-MB-231(FIG. 5B) and PC3 (FIG. 5C) tumor-bearing SCID mice.

FIG. 6 shows the effects of ABI-007 or Taxol used in metronomic or MTDregimes on basic fibroblast growth factor (bFGF)-induced angiogenesis inmatrigel plugs injected subcutaneously into the flanks of Balb/cJ mice.Treatments-A, saline/albumin; B, Cremophor EL control; C, metronomicTaxol 1.3 mg/kg; D, E, and F, metronomic ABI-007 3, 6, and 10 mg/kg,respectively; G, MTD Taxol; H, MTD ABI-007. Matrigel implanted withoutbFGF (−bFGF) served as negative control. Bars, mean±SE.

FIG. 7A and FIG. 7B show the cytotoxic activity of nab-rapamycin incombination with Abraxane™ on vascular smooth muscle cells. Cytotoxicitywas evaluated by staining with ethidium homodimer-1 (FIG. 7A) or bystaining with calcein (FIG. 7B).

FIG. 8 shows the cytotoxic activity of nab-rapamycin in combination withAbraxane™ in a HT29 human colon carcinoma xenograft model.

FIG. 9 shows the cytotoxic activity of nab-17-AAG in combination withAbraxane™ in a H358 human lung carcinoma xenograft model.

DETAILED DESCRIPTION OF THE INVENTION

The present invention provides methods of combination therapy comprisinga first therapy comprising administration of nanoparticles comprising ataxane and a carrier protein (such as albumin) in conjunction with asecond therapy such as radiation, surgery, administration of at leastone other chemotherapeutic agent, or combinations thereof. The inventionalso provides methods of metronomic therapy.

The present invention involves the discovery that Abraxane™, due to itssuperior anti-tumor activity and reduced toxicity and side effects, canbe administered in combination with other therapeutic drugs and/ortreatment modalities and can also be used in metronomic chemotherapy.Due to significantly improved safety profiles with compositionscomprising drug/carrier protein nanoparticles (such as Abraxane™), webelieve that combination chemotherapy with such nanoparticlecompositions (such as Abraxane™) is more effective than combinationchemotherapy with other drugs. In addition the use of nanoparticlecomposition (such as Abraxane™) in combination with radiation is alsobelieved to be more effective than combination of other agents withradiation. Thus, the nanoparticle compositions (especially apaclitaxel/albumin nanoparticle composition, such as Abraxane™), whenused in combination with other chemotherapeutic agents or when combinedwith other treatment modalities, should be very effective and overcomethe deficiencies of surgery, radiation treatment, and chemotherapy inthe treatment of proliferative disease (such as cancer).

The present invention in one its embodiments is the use of a firsttherapy comprising a taxane, such as Abraxane™, in combination with asecond therapy such as another chemotherapeutic agent or agents,radiation, or the like for treating proliferative diseases such ascancer. The first therapy comprising a taxane and second therapy can beadministered to a mammal having the proliferative sequentially, or theycan be co-administered, and even administered simultaneously in the samepharmaceutical composition.

Further, a metronomic dosing regime using Abraxane™ has been found to bemore effective than the traditional MTD dosing schedule of the same drugcomposition. Such metronomic dosing regime of Abraxane™ has also beenfound to be more effective than metronomic dosing of Taxol®.

The methods described herein are generally useful for treatment ofdiseases, particularly proliferative diseases. As used herein,“treatment” is an approach for obtaining beneficial or desired clinicalresults. For purposes of this invention, beneficial or desired clinicalresults include, but are not limited to, any one or more of: alleviationof one or more symptoms, diminishment of extent of disease, stabilized(i.e., not worsening) state of disease, preventing or delaying spread(e.g., metastasis) of disease, preventing or delaying occurrence orrecurrence of disease, delay or slowing of disease progression,amelioration of the disease state, and remission (whether partial ortotal). Also encompassed by “treatment” is a reduction of pathologicalconsequence of a proliferative disease. The methods of the inventioncontemplate any one or more of these aspects of treatment.

As used herein, a “proliferative disease” is defined as a tumor disease(including benign or cancerous) and/or any metastases, wherever thetumor or the metastasis are located, more especially a tumor selectedfrom the group comprising one or more of (and in some embodimentsselected from the group consisting of) breast cancer, genitourinarycancer, lung cancer, gastrointestinal cancer, epidermoid cancer,melanoma, ovarian cancer, pancreatic cancer, neuroblastoma, colorectalcancer, head and neck cancer. In a broader sense of the invention, aproliferative disease may furthermore be selected fromhyperproliferative conditions such as hyperplasias, fibrosis (especiallypulmonary, but also other types of fibrosis, such as renal fibrosis),angiogenesis, psoriasis, atherosclerosis and smooth muscle proliferationin the blood vessels, such as stenosis or restenosis followingangioplasty. In some embodiments, the proliferative disease is cancer.In some embodiments, the proliferative disease is a non-cancerousdisease. In some embodiments, the proliferative disease is a benign ormalignant tumor. Where hereinbefore and subsequently a tumor, a tumordisease, a carcinoma or a cancer are mentioned, also metastasis in theoriginal organ or tissue and/or in any other location are impliedalternatively or in addition, whatever the location of the tumor and/ormetastasis is.

The term “effective, amount” used herein refers to an amount of acompound or composition sufficient to treat a specified disorder,condition or disease such as ameliorate, palliate, lessen, and/or delayone or more of its symptoms. In reference to cancers or other unwantedcell proliferation, an effective amount comprises an amount sufficientto cause a tumor to shrink and/or to decrease the growth rate of thetumor (such as to suppress tumor growth) or to prevent or delay otherunwanted cell proliferation. In some embodiments, an effective amount isan amount sufficient to delay development. In some embodiments, aneffective amount is an amount sufficient to prevent or delay occurrenceand/or recurrence. An effective amount can be administered in one ormore administrations. In the case of cancer, the effective amount of thedrug or composition may: (i) reduce the number of cancer cells; (ii)reduce tumor size; (iii) inhibit, retard, slow to some extent andpreferably stop cancer cell infiltration into peripheral organs; (iv)inhibit (i.e., slow to some extent and preferably stop) tumormetastasis; (v) inhibit tumor growth; (vi) prevent or delay occurrenceand/or recurrence of tumor; and/or (vii) relieve to some extent one ormore of the symptoms associated with the cancer.

In some embodiments, there is provided a method of treating a primarytumor. In some embodiments, there is provided a method of treatingmetastatic cancer (that is, cancer that has metastasized from theprimary tumor). In some embodiments, there is provided a method oftreating cancer at advanced stage(s). In some embodiments, there isprovided a method of treating breast cancer (which may be HER2 positiveor HER2 negative), including, for example, advanced breast cancer, stageIV breast cancer, locally advanced breast cancer, and metastatic breastcancer. In some embodiments, there is provided a method of treating lungcancer, including, for example, non-small cell lung cancer (NSCLC, suchas advanced NSCLC), small cell lung cancer (SCLC, such as advancedSCLC), and advanced solid tumor malignancy in the lung. In someembodiments, there is provided a method of treating any of ovariancancer, head and neck cancer, gastric malignancies, melanoma (includingmetastatic melanoma), colorectal cancer, pancreatic cancer, and solidtumors (such as advanced solid tumors). In some embodiments, there isprovided a method of reducing cell proliferation and/or cell migration.In some embodiments, there is provided a method of treating any of thefollowing diseases: restenosis, stenosis, fibrosis, angiogenesis,psoriasis, atherosclerosis, and proliferation of smooth muscle cells.The present invention also provides methods of delaying development ofany of the proliferative diseases described herein.

The term “individual” is a mammal, including humans. An individualincludes, but is not limited to, human, bovine, horse, feline, canine,rodent, or primate. In some embodiments, the individual is human. Theindividual (such as human) may have advanced disease or lesser extent ofdisease, such as low tumor burden. In some embodiments, the individualis at an early stage of a proliferative disease (such as cancer). Insome embodiments, the individual is at an advanced stage of aproliferative disease (such as an advanced cancer). In some embodiments,the individual is HER2 positive. In some embodiments, the individual isHER2 negative.

The methods may be practiced in an adjuvant setting. “Adjuvant setting”refers to a clinical setting in which an individual has had a history ofa proliferative disease, particularly cancer, and generally (but notnecessarily) been responsive to therapy, which includes, but is notlimited to, surgery (such as surgical resection), radiotherapy, andchemotherapy. However, because of their history of the proliferativedisease (such as cancer), these individuals are considered at risk ofdevelopment of the disease. Treatment or administration in the “adjuvantsetting” refers to a subsequent mode of treatment. The degree of risk(i.e., when an individual in the adjuvant setting is considered as “highrisk” or “low risk”) depends upon several factors, most usually theextent of disease when first treated. The methods provided herein mayalso be practiced in a neoadjuvant setting, i.e., the method may becarried out before the primary/definitive therapy. In some embodiments,the individual has previously been treated. In some embodiments, theindividual has not previously been treated. In some embodiments, thetreatment is a first line therapy.

It is understood that aspect and embodiments of the invention describedherein include “consisting” and/or “consisting essentially of” aspectsand embodiments.

Combination Therapy with Chemotherapeutic Agent

The present invention provides methods of treating a proliferativedisease (such as cancer) in an individual, comprising administering tothe individual: a) an effective amount of a composition comprisingnanoparticles comprising a taxane and a carrier protein (such asalbumin); and b) an effective amount of at least one otherchemotherapeutic agent. In some embodiments, the taxane is any of (andin come embodiments consisting essentially of) paclitaxel, docetaxel,and ortataxel. In some embodiments, the nanoparticle compositioncomprises Abraxane™. In some embodiments, the chemotherapeutic agent isany of (and in some embodiments selected from the group consisting of)antimetabolite agents (including nucleoside analogs), platinum-basedagents, alkylating agents, tyrosine kinase inhibitors, anthracyclineantibiotics, vinca alkloids, proteasome inhibitors, macrolides, andtopoisomerase inhibitors.

In some embodiments, the method comprises administering to theindividual: a) an effective amount of a composition comprisingnanoparticles comprising paclitaxel and an albumin; and b) an effectiveamount of at least one other chemotherapeutic agent. In someembodiments, the paclitaxel/albumin nanoparticles have an averagediameter of no greater than about 200 nm. In some embodiments, thepaclitaxel/albumin nanoparticle composition is substantially free (suchas free) of surfactant (such as Cremophor). In some embodiments, theweight ratio of the albumin to paclitaxel in the composition is about18:1 or less, such as about 9:1 or less. In some embodiments, thepaclitaxel is coated with albumin. In some embodiments, thepaclitaxel/albumin nanoparticles have an average diameter of no greaterthan about 200 nm and the paclitaxel/albumin composition issubstantially free (such as free) of surfactant (such as Cremophor). Insome embodiments, the paclitaxel/albumin nanoparticles have an averagediameter of no greater than about 200 nm and the paclitaxel is coatedwith albumin. In some embodiments, the nanoparticle composition isAbraxane™.

In some embodiments, the invention provides a method of treating aproliferative disease (such as cancer) in an individual comprisingadministering to the individual a) an effective amount of Abraxane™, andb) an effective amount of at least one other chemotherapeutic agent.Preferred drug combinations for sequential or co-administration orsimultaneous administration with Abraxane™ are those which show enhancedantiproliferative activity when compared with the single componentsalone, especially combinations that that lead to regression ofproliferative tissues and/or cure from proliferative diseases.

The chemotherapeutic agents described herein can be the agentsthemselves, pharmaceutically acceptable salts thereof, andpharmaceutically acceptable esters thereof, as well as steroisomers,enantiomers, racemic mixtures, and the like. The chemotherapeutic agentor agents as described can be administered as well as a pharmaceuticalcomposition containing the agent(s), wherein the pharmaceuticalcomposition comprises a pharmaceutically acceptable-carrier vehicle, orthe like.

The chemotherapeutic agent may be present in a nanoparticle composition.For example, in some embodiments, there is provided a method of treatinga proliferative disease (such as cancer) in an individual, comprisingadministering to the individual: a) an effective amount of a compositioncomprising nanoparticles comprising a taxane and a carrier protein (suchas albumin); and b) an effective amount of a composition comprisingnanoparticles comprising at least one other chemotherapeutic agent and acarrier protein (such as albumin). In some embodiments, the methodcomprises administering to the individual: a) an effective amount of acomposition comprising nanoparticles comprising paclitaxel and analbumin (such as Abraxane™); and b) an effective amount of a compositioncomprising nanoparticles comprising at least one other chemotherapeuticagent and a carrier protein (such as albumin). In some embodiments, thechemotherapeutic agent is any of (and in some embodiments selected fromthe group consisting of) thiocolchicine or its derivatives (such asdimeric thiocolchicine, including for example nab-5404, nab-5800, andnab-5801), rapamycin or its derivatives, and geldanamycin or itsderivatives (such as 17-allyl amino geldanamycin (17-AAG)). In someembodiments, the chemotherapeutic agent is rapamycin. In someembodiments, the chemotherapeutic agent is 17-AAG.

An exemplary and non-limiting list of chemotherapeutic agentscontemplated is provided herein. Suitable chemotherapeutic agentsinclude, for example, vinca alkaloids, agents that disrupt microtubuleformation (such as colchicines and its derivatives), anti-angiogenicagents, therapeutic antibodies, EGFR targeting agents, tyrosine kinasetargeting agent (such as tyrosine kinase inhibitors), transitional metalcomplexes, proteasome inhibitors, antimetabolites (such as nucleosideanalogs), alkylating agents, platinum-based agents, anthracyclineantibiotics, topoisomerase inhibitors, macrolides, therapeuticantibodies, retinoids ( such as all-trans retinoic acids or aderivatives thereof); geldanamycin or a derivative thereof (such as17-AAG), and other standard chemotherapeutic agents well recognized inthe art.

In some embodiments, the chemotherapeutic agent is any of (and in someembodiments selected from the group consisting of) adriamycin,colchicine, cyclophosphamide, actinomycin, bleomycin, duanorubicin,doxorubicin, epirubicin, mitomycin, methotrexate, mitoxantrone,fluorouracil, carboplatin, carmustine (BCNU), methyl-CCNU, cisplatin,etoposide, interferons, camptothecin and derivatives thereof,phenesterine, taxanes and derivatives thereof (e.g., paclitaxel andderivatives thereof, taxotere and derivatives thereof, and the like),topetecan, vinblastine, vincristine, tamoxifen, piposulfan, nab-5404,nab-5800, nab-5801, Irinotecan, HKP, Ortataxel, gemcitabine, Herceptin®,vinorelbine, Doxili®, capecitabine, Alimta®, Avastin®, Velcade®,Tarceva®, Neulasta®, Lapatinib, Sorafenib, derivatives thereof,chemotherapeutic agents known in the art, and the like. In someembodiments, the chemotherapeutic agent is a composition comprisingnanoparticles comprising a thiocolchicine derivative and a carrierprotein (such as albumin).

In some embodiments, the chemotherapeutic agent is a antineoplasticagent including, but is not limited to, carboplatin, Navelbine®(vinorelbine), anthracycline (Doxil®), lapatinib (GW57016), Herceptin®,gemcitabine (Gemzar®), capecitabine (Xeloda®), Alimta®, cisplatin,5-fluorouracil, epirubicin, cyclophosphamide, Avastin®, Velcade®, etc.

In some embodiments, the chemotherapeutic agent is an antagonist ofother factors that are involved in tumor growth, such as EGFR, ErbB2(also known as Herb), ErbB3, ErbB4, or TNF. Sometimes, it may bebeneficial to also administer one or more cytokines to the individual.In some embodiments, the therapeutic agent is a growth inhibitory agent.Suitable dosages for the growth inhibitory agent are those presentlyused and may be lowered due to the combined action (synergy) of thegrowth inhibitory agent and the taxane.

In some embodiments, the chemotherapeutic agent is a chemotherapeuticagent other than an anti-VEGF antibody, a HER2 antibody, interferon, andan HGFβ antagonist.

Reference to a chemotherapeutic agent herein applies to thechemotherapeutic agent or its derivatives and accordingly the inventioncontemplates and includes either of these embodiments (agent; agent orderivative(s)). “Derivatives” or “analogs” of a chemotherapeutic agentor other chemical moiety include, but are not limited to, compounds thatare structurally similar to the chemotherapeutic agent or moiety or arein the same general chemical class as the chemotherapeutic agent ormoiety. In some embodiments, the derivative or analog of thechemotherapeutic agent or moiety retains similar chemical and/orphysical property (including, for example, functionality) of thechemotherapeutic agent or moiety.

In some embodiments, the invention provides a method of treating aproliferative disease (such as cancer) in an individual, comprisingadministering to the individual a) an effective amount of a compositioncomprising nanoparticles comprising a taxane and a carrier protein (suchas albumin), and b) an effective amount of a tyrosine kinase inhibitor.In some embodiments, the invention provides a method of treating aproliferative disease (such as cancer) in an individual, comprisingadministering to the individual a) an effective amount of a compositioncomprising nanoparticles comprising paclitaxel and an albumin (such asAbraxane™), and b) an effective amount of a tyrosine kinase inhibitor.Suitable tyrosine kinase inhibitors include, for example, imatinib(Gleevec®), gefitinib (Iressa®), Tarceva, Sutent® (sunitinib malate),and Lapatinib. In some embodiments, the tyrosine kinase inhibitor islapatinib. In some embodiments, the tyrosine kinase inhibitor isTarceva. Tarceva is a small molecule human epidermal growth factor type1/epidermal growth factor receptor (HER1/EGFR) inhibitor whichdemonstrated, in a Phase III clinical trial, an increased survival inadvanced non-small cell lung cancer (NSCLC) individuals. In someembodiments, the method is for treatment of breast cancer, includingtreatment of metastatic breast cancer and treatment of breast cancer ina neoadjuvant setting. In some embodiments, the method is for treatmentof advanced solid tumor. In some embodiments, there is provided a methodto inhibit the proliferation of EGFR expressing tumors in a mammalcomprising administering to a mammal infected with such tumors Abraxane™and gefitinib, wherein the gefitinib is administered by pulse-dosing.

In some embodiments, the invention provides a method of treating aproliferative disease (such as cancer) in an individual, comprisingadministering to the individual a) an effective amount of a compositioncomprising nanoparticles comprising a taxane and a carrier protein (suchas albumin), and b) an effective amount of an antimetabolite agent (suchas a nucleoside analog, including for example purine analogs andpyrimidine analogs). In some embodiments, the invention provides amethod of treating a proliferative disease (such as cancer) in anindividual, comprising administering to the individual a) an effectiveamount of a composition comprising nanoparticles comprising paclitaxeland an albumin (such as Abraxane™), and b) an effective amount of anantimetabolite agent. An “antimetabolic agent” is an agent which isstructurally similar to a metabolite, but cannot be used by the body ina productive manner. Many antimetabolite agents interfere withproduction of nucleic acids, RNA and DNA. For example, theantimetabolite can be a nucleoside analog, which includes, but is notlimited to, azacitidine, azathioprine, capecitabine (Xeloda®),cytarabine, cladribine, cytosine arabinoside (ara-C, cytosar),doxifluridine, fluorouracil (such as 5-fluorouracil), UFT, hydoxyurea,gemcitabine, mercaptopurine, methotrexate, thioguanine (such as6-thioguanine). Other anti-metabolites include, for example,L-asparaginase (Elspa), decarbazine (DTIC), 2-deoxy-D-glucose, andprocarbazine (matulane). In some embodiments, the nucleoside analog isany of (and in some embodiments selected from the group consisting of)gemcitabine, fluorouracil, and capecitabine. In some embodiments, themethod is for treatment of metastatic breast cancer or locally advancedbreast cancer. In some embodiments, the method is for first linetreatment of metastatic breast cancer. In some embodiments, the methodis for treatment of breast cancer in a neoadjuvant setting. In someembodiments, the method is for treatment of any of NSCLC, metastaticcolorectal cancer, pancreatic cancer, or advanced solid tumor.

In some embodiments, the invention provides a method of treating aproliferative disease (such as cancer) in an individual, comprisingadministering to the individual a) an effective amount of a compositioncomprising nanoparticles comprising a taxane and a carrier protein (suchas albumin), and b) an effective amount of an alkylating agent. In someembodiments, the invention provides a method of treating a proliferativedisease (such as cancer) in an individual, comprising administering tothe individual a) an effective amount of a composition comprisingnanoparticles comprising paclitaxel and an albumin (such as Abraxane™),and b) an effective amount of an alkylating agent. Suitable alkylatingagents include, but are not limited to, cyclophosphamide (Cytoxan),mechlorethamine, chlorambucil, melphalan, carmustine (BCNU), thiotepa,busulfan, alkyl sulphonates, ethylene imines, nitrogen mustard analogs,estramustine sodium phosphate, ifosfamide, nitrosoureas, lomustine, andstreptozocin. In some embodiments, the alkylating agent iscyclophosphamide. In some embodiments, the cyclophosphamide isadministered prior to the administration of the nanoparticlecomposition. In some embodiments, the method is for treatment of anearly stage breast cancer. In some embodiments, the method is fortreatment of a breast cancer in an adjuvant or a neoadjuvant setting.

In some embodiments, the invention provides a method of treating aproliferative disease (such as cancer) in an individual, comprisingadministering to the individual a) an effective amount of a compositioncomprising nanoparticles comprising a taxane and a carrier protein (suchas albumin), and b) an effective amount of a platinum-based agent. Insome embodiments, the invention provides a method of treating aproliferative disease (such as cancer) in an individual, comprisingadministering to the individual a) an effective amount of a compositioncomprising nanoparticles comprising paclitaxel and an albumin (such asAbraxane™), and b) an effective amount of a platinum-based agent.Suitable platinum-based agents include, but are not limited to,carboplatin, cisplatin, and oxaliplatin. In some embodiments, theplatinum-based agent is carboplatin. In some embodiments, the method isfor treatment of: breast cancer (HER2 positive or HER2 negative,including metastatic breast cancer and advanced breast cancer); lungcancer (including advanced NSCLC, first line NSCLC, SCLC, and advancedsolid tumor malignancies in the lung); ovarian cancer; head and neckcancer; and melanoma (including metastatic melanoma).

In some embodiments, the invention provides a method of treating aproliferative disease (such as cancer) in an individual, comprisingadministering to the individual a) an effective amount of a compositioncomprising nanoparticles comprising a taxane and a carrier protein (suchas albumin), and b) an effective amount of an anthracycline antibiotic.In some embodiments, the invention provides a method of treating aproliferative disease (such as cancer) in an individual, comprisingadministering to the individual a) an effective amount of a compositioncomprising nanoparticles comprising paclitaxel and an albumin (such asAbraxane™) and a carrier protein (such as albumin), and b) an effectiveamount of an anthracycline antibiotic. Suitable anthracycline antibioticinclude, but are not limited to, Doxil®, actinomycin, dactinomycin,daunorubicin (daunomycin), doxorubicin (adriamycin), epirubicin,idarubicin, mitoxantrone, valrubicin. In some embodiments, theanthracycline is any of (and in some embodiments selected from the groupconsisting of) Doxil®, epirubicin, and doxorubicin. In some embodiments,the method is for treatment of an early stage breast cancer. In someembodiments, the method is for treatment of a breast cancer in anadjuvant or a neoadjuvant setting.

In some embodiments, the invention provides a method of treating aproliferative disease (such as cancer) in an individual, comprisingadministering to the individual a) an effective amount of a compositioncomprising nanoparticles comprising a taxane and a carrier protein (suchas albumin), and b) an effective amount of a vinca alkloid. In someembodiments, the invention provides a method of treating a proliferativedisease (such as cancer) in an individual, comprising administering tothe individual a) an effective amount of a composition comprisingnanoparticles comprising palitaxel and an albumin (such as Abraxane™)and a carrier protein (such as albumin), and b) an effective amount of avinca alkloid. Suitable vinca alkaloids include, for example,vinblastine, vincristine, vindesine, vinorelbine (Navelbine®), andVP-16. In some embodiments, the vinca alkaloid is vinorelbine(Navelbine®). In some embodiments, the method is for treatment of stageIV breast cancer and lung cancer.

In some embodiments, the invention provides a method of treating aproliferative disease (such as cancer) in an individual, comprisingadministering to the individual a) an effective amount of a compositioncomprising nanoparticles comprising a taxane and a carrier protein (suchas albumin), and b) an effective amount of a macrolide. In someembodiments, the invention provides a method of treating a proliferativedisease (such as cancer) in an individual, comprising administering tothe individual a) an effective amount of a composition comprisingnanoparticles comprising paclitaxel and an albumin (such as Abraxane™)and a carrier protein (such as albumin); and b) an effective amount of amacrolide. Suitable macrolides include, for example, rapamycin,carbomycin, and erythromycin. In some embodiments, the macrolide israpamycin or a derivative thereof. In some embodiments, the method isfor treatment of a solid tumor.

In some embodiments, the invention provides a method of treating aproliferative disease (such as cancer) in an individual, comprisingadministering to the individual a) an effective amount of a compositioncomprising nanoparticles comprising a taxane and a carrier protein (suchas albumin), and b) an effective amount of a topoisomerase inhibitor. Insome embodiments, the invention provides a method of treating aproliferative disease (such as cancer) in an individual, comprisingadministering to the individual a) an effective amount of a compositioncomprising nanoparticles comprising paclitaxel and an albumin (such asAbraxane™) and a carrier protein (such as albumin), and b) an effectiveamount of a topoisomerase inhibitor. In some embodiments, thechemotherapeutic agent is a topoisomerase inhibitor, including, forexample, inhibitor of topoisomerase I and topoisomerase II. Exemplaryinhibitors of topoisomerase I include, but are not limited to,camptothecin, such as irinotecan and topotecan. Exemplary inhibitors oftopoisomerase II include, but are not limited to, amsacrine, etoposide,etoposide phosphate, and teniposide.

In some embodiments, the invention provides a method of treating aproliferative disease (such as cancer) in an individual, comprisingadministering to the individual a) an effective amount of a compositioncomprising nanoparticles comprising a taxane and a carrier protein (suchas albumin), and b) an effective amount of an antiangiogenic agent. Insome embodiments, the invention provides a method of treating aproliferative disease (such as cancer) in an individual, comprisingadministering to the individual a) an effective amount of a compositioncomprising nanoparticles comprising paclitaxel and an albumin (such asAbraxane™) and a carrier protein (such as albumin), and b) an effectiveamount of an antiangiogenic agent. In some embodiments, the method isfor treatment of metastatic breast cancer, breast cancer in an adjuvantsetting or a neoadjuvant setting, lung cancer (such as first lineadvanced NSCLC and NSCLC), ovarian cancer, and melanoma (includingmetastatic melanoma).

Many anti-angiogenic agents have been identified and are known in theart, including those listed by Carmeliet and Jain (2000). Theanti-angiogenic agent can be naturally occurring or non-naturallyoccurring. In some embodiments, the chemotherapeutic agent is asynthetic antiangiogenic peptide. For example, it has been previouslyreported that the antiangiogenic activity of small synthetic pro-apopticpeptides comprise two functional domains, one targeting the CD13receptors (aminopeptidase N) on tumor microvessels and the otherdisrupting the mitochondrial membrane following internalization. Nat.Med. 1999, 5(9):1032-8. A second generation dimeric peptide,CNGRC-GG-d(KLAKLAK)2, named HKP (Hunter Killer Peptide) was found tohave improved antitumor activity. Accordingly, in some embodiments, theantiangiogenic peptide is HKP. In some embodiments, the antiangiogenicagent is other than an anti-VEGF antibody (such as Avastin®).

In some embodiments, the invention provides a method of treating aproliferative disease (such as cancer) in an individual, comprisingadministering to the individual a) an effective amount of a compositioncomprising nanoparticles comprising a taxane and a carrier protein (suchas albumin), and b) an effective amount of a proteasome inhibitor, suchas bortezomib.(Velcade). In some embodiments, the invention provides amethod of treating a proliferative disease (such as cancer) in anindividual, comprising administering to the individual a) an effectiveamount of a composition comprising nanoparticles comprising paclitaxeland an albumin (such as Abraxane™) and a carrier protein (such asalbumin), and b) an effective amount of a proteasome inhibitor such asbortezomib (Velcade).

In some embodiments, the invention provides a method of treating aproliferative disease (such as cancer) in an individual, comprisingadministering to the individual a) an effective amount of a compositioncomprising nanoparticles comprising a taxane and a carrier protein (suchas albumin), and b) an effective amount of a therapeutic antibody. Insome embodiments, the invention provides a method of treating aproliferative disease (such as cancer) in an individual, comprisingadministering to the individual a) an effective amount of a compositioncomprising nanoparticles comprising paclitaxel and an albumin (such asAbraxane™) and a carrier protein (such as albumin), and b) an effectiveamount of a therapeutic antibody. Suitable therapeutic antibodiesinclude, but are not limited to, anti-VEGF antibody (such as Avastin®(bevacizumab)), anti-HER2 antibody (such as Herceptin® (trastuzumab)),Erbitux® (cetuximab), Campath (alemtuzumab), Myelotarg (gemtuzumab),Zevalin (ibritumomab tiuextan, Rituxan (rituximab), and Bexxar(tositumomab). In some embodiments, the chemotherapeutic agent isErbitux® (cetuximab). In some embodiments, the chemotherapeutic agent isa therapeutic antibody other than an antibody against VEGF or HER2. Insome embodiments, the method is for treatment of HER2 positive breastcancer, including treatment of advanced breast cancer, treatment ofmetastatic cancer, treatment of breast cancer in an adjuvant setting,and treatment of cancer in a neoadjuvant setting. In some embodiments,the method is for treatment of any of metastatic breast cancer, breastcancer in an adjuvant setting or a neoadjuvant setting, lung cancer(such as first line advanced NSCLC and NSCLC), ovarian cancer, head andneck cancer, and melanoma (including metastatic melanoma). For example,in some embodiments, there is provided a method for treatment of HER2positive metastatic breast cancer in an individual, comprisingadministering to the individual 125 mg/m² paclitaxel/albuminnanoparticle composition (such as Abraxane™) weekly for three weeks withthe fourth week off, concurrent with the administration of Herceptin®.

In some embodiments, two or more chemotherapeutic agents areadministered in addition to the taxane in the nanoparticle composition.These two or more chemotherapeutic agents may (but not necessarily)belong to different classes of chemotherapeutic agents. Examples ofthese combinations are provided herein. Other combinations are alsocontemplated.

In some embodiments, there is provided a method of treating aproliferative disease (such as cancer) in an individual, comprisingadministering to the individual a) an effective amount of a compositioncomprising nanoparticles comprising a taxane and a carrier protein (suchas albumin), b) an effective amount of an antimetabolite (such as anucleoside analog, for example, gemcitabine), and c) an anthracyclineantibiotic (such as epirubicin). In some embodiments, there is provideda method of treating a proliferative disease (such as cancer) in anindividual, comprising administering to the individual a) an effectiveamount of a composition comprising nanoparticles comprising paclitaxeland an albumin (such as Abraxane™), b) an effective amount of anantimetabolite (such as a nucleoside analog, for example, gemcitabine),and c) an effective amount of an anthracycline antibiotic (such asepirubicin). In some embodiments, the method is for treatment of breastcancer in a neoadjuvant setting. For example, in some embodiments, thereis provided a method of treating locally advanced/inflammatory cancer inan individual comprising administering to the individual 220 mg/m²paclitaxel/albumin nanoparticle composition (such as Abraxane™) everytwo weeks; 2000 mg/m² gemcitabine, every two weeks; and 50 mg/m²epirubicin, every two weeks. In some embodiments, there is provided amethod of treating breast cancer in an individual in an adjuvantsetting, comprising administering to the individual 175 mg/m²paclitaxel/albumin nanoparticle composition (such as Abraxane™) everytwo weeks, 2000 mg/m² gemcitabine, every two weeks, and 50 mg/m²epirubicin, every two weeks.

In some embodiments, there is provided a method of treating aproliferative disease (such as cancer) in an individual, comprisingadministering to the individual a) an effective amount of a compositioncomprising nanoparticles comprising a taxane and a carrier protein (suchas albumin), b) an effective amount of a platinum-based agent (such ascarboplatin), and c) a therapeutic antibody (such as ant-HER2 antibody(such as Herceptin®) and anti-VEGF antibody (such as Avastin®)). In someembodiments, there is provided a method of treating a proliferativedisease (such as cancer) in an individual, comprising administering tothe individual a) an effective amount of a composition comprisingnanoparticles comprising paclitaxel and an albumin (such as Abraxane™),b) an effective amount of a platinum-based agent (such as carboplatin),and c) a therapeutic antibody (such as ant-HER2 antibody (such asHerceptin®l and anti-VEGF antibody (such as Avastin®)). In someembodiments, the method is for treatment of any of advanced breastcancer, metastatic breast cancer, breast cancer in an adjuvant setting,and lung cancer (including NSCLC and advanced NSCLC). In someembodiments, there is provided a method of treating metastatic cancer inan individual, comprising administering to the individual 75 mg/m²paclitaxel/albumin nanoparticle composition (such as Abraxane™) andcarboplatin, AUC=2, wherein the administration is carried out weekly forthree weeks with the fourth week off. In some embodiments, the methodfurther comprises weekly administering about 2-4 mg/kg of Herception®.

In some embodiments, there is provided a method of treating aproliferative disease (such as cancer) in an individual, comprisingadministering to the individual a) an effective amount of a compositioncomprising nanoparticles comprising a taxane and a carrier protein (suchas albumin), b) an effective amount of a platinum-based agent (such ascarboplatin), and c) a vinca alkaloid (such as Navelbine®). In someembodiments, there is provided a method of treating a proliferativedisease (such as cancer) in an individual, comprising administering tothe individual a) an effective amount of a composition comprisingnanoparticles comprising paclitaxel and an albumin (such as Abraxane™),b) an effective amount of a platinum-based agent (such as carboplatin),and c) a vinca alkaloid (such as Navelbine®). In some embodiments, themethod is for treatment of lung cancer.

In some embodiments, the invention provides a method of treating aproliferative disease (such as cancer) in an individual, comprisingadministering to the individual a) an effective amount of a compositioncomprising nanoparticles comprising a taxane and a carrier protein (suchas albumin), b) an effective amount of an alkylating agent (such ascyclophosphamide) and c) an anthracycline antibiotic (such asadriamycin). In some embodiments, the invention provides a method oftreating a proliferative disease (such as cancer) in an individual,comprising administering to the individual a) an effective amount of acomposition comprising nanoparticles comprising paclitaxel and analbumin, b) an effective amount of an alkylating agent (such ascyclophosphamide) and c) an anthracycline antibiotic (such asadriamycin). In some embodiments, the method is for treatment of anearly stage breast cancer. In some embodiments, the method is fortreatment of a breast cancer in an adjuvant or a neoadjuvant setting.For example, in some embodiments, there is provided a method of treatingan early stage breast cancer in an individual, comprising administering260 mg/m² paclitaxel/albumin nanoparticle composition (such asAbraxane™), 60 mg/m² adriamycin, and 600 mg/m² cyclophosphamide, whereinthe administration is carried out once every two weeks.

Other embodiments are provided in Table 1. For example, in someembodiments, there is provided a method of treating advanced breastcancer in an individual, comprising administering to the individual a)an effective amount of a composition comprising nanoparticles comprisinga paclitaxel and an albumin (such as Abraxane™), b) an effective amountof carboplatin. In some embodiments, the method further comprisesadministering an effective amount of Herceptin® to the individual. Insome embodiments, there is provided a method of treating metastaticbreast cancer in an individual, comprising administering to theindividual a) an effective amount of a composition comprisingnanoparticles comprising paclitaxel and an albumin (such as Abraxane™),b) an effective amount of gemcitabine. In some embodiments, there isprovided a method of treating advanced non-small cell lung cancer in anindividual, comprising administering to the individual a) an effectiveamount of a composition comprising nanoparticles comprising paclitaxeland an albumin (such as Abraxane™), b) an effective amount ofcarboplatin.

In some embodiments, there is provided a composition comprisingnanoparticles comprising a taxane (such as paclitaxel, docetaxel, orortataxel) and a carrier protein (such as albumin) and at least oneother chemotherapeutic agent. The compositions described herein maycomprise effective amounts of the taxane and the chemotherapeutic agentfor the treatment of a proliferative disease (such as cancer). In someembodiments, the chemotherapeutic agent and the taxane are present inthe composition at a predetermined ratio, such as the weight ratiosdescribed herein. In some embodiments, the invention provides asynergistic composition of an effective amount of a compositioncomprising nanoparticles comprising a taxane (such as paclitaxel,docetaxel, or ortataxel) and an effective amount of at least one otherchemotherapeutic agent.

In some embodiments, the invention provides pharmaceutical compositionscomprising nanoparticles comprising a taxane and a carrier protein (suchas albumin) for use in the treatment of a proliferative disease (such ascancer), wherein said use comprises simultaneous and/or sequentialadministration of at least one other chemotherapeutic agent. In someembodiments, the invention provides a pharmaceutical compositioncomprising a chemotherapeutic agent for use in the treatment of aproliferative disease (such as cancer), wherein said use comprisessimultaneous and/or sequential administration of a compositioncomprising nanoparticles comprising a taxane and a carrier protein (suchas albumin). In some embodiments, the invention providestaxane-containing nanoparticle compositions and compositions comprisingone other chemotherapeutic agent for simultaneous, and/or sequential usefor treatment of a proliferative disease (such as cancer).

Modes of Administration

The composition comprising nanoparticles comprising taxane (alsoreferred to as “nanoparticle composition”) and the chemotherapeuticagent can be administered simultaneously (i.e., simultaneousadministration) and/or sequentially (i.e., sequential administration).

In some embodiments, the nanoparticle composition and thechemotherapeutic agent (including the specific chemotherapeutic agentsdescribed herein) are administered simultaneously. The term“simultaneous administration,” as used herein, means that thenanoparticle composition and the chemotherapeutic agent are administeredwith a time separation of no more than about 15 minute(s), such as nomore than about any of 10, 5, or 1 minutes. When the drugs areadministered simultaneously, the drug in the nanoparticles and thechemotherapeutic agent may be contained in the same composition (e.g., acomposition comprising both the nanoparticles and the chemotherapeuticagent) or in separate compositions (e.g., the nanoparticles arecontained in one composition and the chemotherapeutic agent is containedin another composition). For example, the taxane and thechemotherapeutic agent may be present in a single composition containingat least two different nanoparticles, wherein some of the nanoparticlesin the composition comprise the taxane and a carrier protein, and someof the other nanoparticles in the composition comprise thechemotherapeutic agent and a carrier protein. The invention contemplatesand encompasses such compositions. In some embodiments, only the taxaneis contained in nanoparticles. In some embodiments, simultaneousadministration of the drug in the nanoparticle composition and thechemotherapeutic agent can be combined with supplemental doses of thetaxane and/or the chemotherapeutic agent.

In some embodiments, the nanoparticle composition and thechemotherapeutic agent are administered sequentially. The term“sequential administration” as used herein means that the drug in thenanoparticle composition and the chemotherapeutic agent are administeredwith a time separation of more than about 15 minutes, such as more thanabout any of 20, 30, 40, 50,-60 or more minutes. Either the nanoparticlecomposition or the chemotherapeutic agent may be administered first. Thenanoparticle composition and the chemotherapeutic agent are contained inseparate compositions, which may be contained in the same or differentpackages.

In some embodiments, the administration of the nanoparticle compositionand the chemotherapeutic agent are concurrent, i.e., the administrationperiod of the nanoparticle composition and that of the chemotherapeuticagent overlap with each other. In some embodiments, the administrationof the nanoparticle composition and the chemotherapeutic agent arenon-concurrent. For example, in some embodiments, the administration ofthe nanoparticle composition is terminated before the chemotherapeuticagent is administered. In some embodiments, the administration of thechemotherapeutic agent is terminated before the nanoparticle compositionis administered. The time period between these two non-concurrentadministrations can range from about two to eight weeks, such as aboutfour weeks.

The dosing frequency of the drug-containing nanoparticle composition andthe chemotherapeutic agent may be adjusted over the course of thetreatment, based on the judgment of the administering physician. Whenadministered separately, the drug-containing nanoparticle compositionand the chemotherapeutic agent can be administered at different dosingfrequency or intervals. For example, the drug-containing nanoparticlecomposition can be administered weekly, while a chemotherapeutic agentcan be administered more or less frequently. In some embodiments,sustained continuous release formulation of the drug-containingnanoparticle and/or chemotherapeutic agent may be used. Variousformulations and devices for achieving sustained release are known inthe art.

The nanoparticle composition and the chemotherapeutic agent can beadministered using the same route of administration or different routesof administration. In some embodiments (for both simultaneous andsequential administrations), the taxane in the nanoparticle compositionand the chemotherapeutic agent are administered at a predeterminedratio. For example, in some embodiments, the ratio by weight of thetaxane in the nanoparticle composition and the chemotherapeutic agent isabout 1 to 1. In some embodiments, the weight ratio may be between about0.001 to about 1 and about 1000 to about 1, or between about 0.01 toabout 1 and 100 to about 1. In some embodiments, the ratio by weight ofthe taxane in the nanoparticle composition and the chemotherapeuticagent is less than about any of 100:1, 50:1, 30:1, 10:1, 9:1, 8:1, 7:1,6:1, 5:1, 4:1, 3:1, 2: 1, and 1:1 In some embodiments, the ratio byweight of the taxane in the nanoparticle composition and thechemotherapeutic agent is more than about any of 1:1, 2:1, 3:1, 4:1,5:1, 6:1, 7:1, 8:1, 9:1, 30:1, 50:1, 100:1. Other ratios arecontemplated.

The doses required for the taxane and/or the chemotherapeutic agent may(but not necessarily) be lower than what is normally required when eachagent is administered alone. Thus, in some embodiments, a subtherapeuticamount of the drug in the nanoparticle composition and/or thechemotherapeutic agent are administered. “Subtherapeutic amount” or“subtherapeutic level” refer to an amount that is less than thetherapeutic amount, that is, less than the amount normally used when thedrug in the nanoparticle composition and/or the chemotherapeutic agentare administered alone. The reduction may be reflected in terms of theamount administered at a given administration and/or the amountadministered over a given period of time (reduced frequency).

In some embodiments, enough chemotherapeutic agent is administered so asto allow reduction of the normal dose of the drug in the nanoparticlecomposition required to effect the same degree of treatment by at leastabout any of 5%, 10%, 20%, 30%, 50%, 60%, 70%, 80%, 90%, or more. Insome embodiments, enough drug in the nanoparticle composition isadministered so as to allow reduction of the normal, dose of thechemotherapeutic agent required to effect the same degree of treatmentby at least about any of 5%, 10%, 200%o, 30%, 50%, 60%, 70%, 80%, 90%,or more.

In some embodiments, the dose of both the taxane in the nanoparticlecomposition and the chemotherapeutic agent are reduced as compared tothe corresponding normal dose of each when administered alone. In someembodiments, both the taxane in the nanoparticle composition and thechemotherapeutic agent are administered at a subtherapeutic, i.e.,reduced, level. In some embodiments, the dose of the nanoparticlecomposition and/or the chemotherapeutic agent is substantially less thanthe established maximum toxic dose (MTD). For example, the dose of thenanoparticle composition and/or the chemotherapeutic agent is less thanabout 50%, 40%, 30%, 20%, or 10% of the MTD.

A combination of the administration configurations described herein canbe used. The combination therapy methods described herein may beperformed alone or in conjunction with another therapy, such as surgery,radiation, chemotherapy, immunotherapy, gene therapy, and the like.Additionally, a person having a greater risk of developing theproliferative disease may receive treatments to inhibit or and/or delaythe development of the disease.

As will be understood by those of ordinary skill in the art, theappropriate doses of chemotherapeutic agents will be approximately thosealready employed in clinical therapies wherein the chemotherapeuticagent are administered alone or in combination with otherchemotherapeutic agents. Variation in dosage will likely occur dependingon the condition being treated. As described above, in some embodiments,the chemotherapeutic agents may be administered at a reduced level.

The nanoparticle compositions described herein can be administered to anindividual (such as human) via various routes, such as parenterally,including intravenous, intra-arterial, intraperitoneal, intrapulmonary,oral, inhalation, intravesicular, intramuscular, intra-tracheal,subcutaneous, intraocular, intrathecal, or transdermal. For example, thenanoparticle composition can be administered by inhalation to treatconditions of the respiratory tract. The composition can be used totreat respiratory conditions such as pulmonary fibrosis, broncheolitisobliterans, lung cancer, bronchoalveolar carcinoma, and the like. Insome embodiments, the nanoparticle composition is administratedintravenously. In some embodiments, the nanoparticle composition isadministered orally.

The dosing frequency of the administration of the nanoparticlecomposition depends on the nature of the combination therapy and theparticular disease being treated. An exemplary dosing frequency include,but is not limited to, weekly without break; weekly, three out of fourweeks; once every three weeks; once every two weeks; weekly, two out ofthree weeks. See also Table 1.

The dose of the taxane in the nanoparticle composition will vary withthe nature of the combination therapy and the particular disease beingtreated. The dose should be sufficient to effect a desirable response,such as a therapeutic or prophylactic response against a particulardisease. An exemplary dose of the taxane (in some embodimentspaclitaxel) in the nanoparticle composition include, but is not limitedto, about any of 50 mg/m², 60 mg/m², 75 mg/m², 80 mg/m², 90 mg/m², 100mg/m², 120 mg/m², 160 mg/m², 175 mg/M², 200 mg/M², 210 mg/M², 220 mg/m²,260 mg/m², and 300 mg/m². For example, the dosage of paclitaxel in ananoparticle composition can be in the range of 100-400 mg/m² when givenon a 3 week schedule, or 50-250 mg/m² when given on a weekly schedule.See also Table 1.

Other exemplary dosing schedules for the administration of thenanoparticle composition (such as paclitaxel/albumin nanoparticlecomposition, for example Abraxane™) include, but are not limited to, 100mg/m², weekly, without break; 75 mg/m² weekly, 3 out of four weeks; 100mg/m², weekly, 3 out of 4 weeks; 125 mg/m², weekly, 3 out of 4 weeks;125 mg/m², weekly, 2 out of 3 weeks; 130 Mg/m², weekly, without break;175 mg/m², once every 2 weeks; 260 mg/m², once every 2 weeks; 260 mg/m²,once every 3 weeks; 180-300 mg/m², every three weeks; 60-175 mg/m²,weekly, without break. In addition, the taxane (alone or in combinationtherapy) can be administered by following a metronomic dosing regimedescribed herein.

Exemplary dosing regimes for the combination therapy of nanoparticlecomposition (such as paclitaxel/albumin nanoparticle composition, forexample Abraxane™) and other agents include, but are not limited to, 125mg/m² weekly, two out of three weeks, plus 825 mg/m² Xeloda®, daily; 260mg/m² once every two weeks, plus 60 mg/m² adriamycin and 600 mg/m²cyclophosphamide, once every two weeks; 220-340 mg/m² once every threeweeks, plus carboplatin, AUC=6, once every three weeks; 100-150 mg/m²weekly, plus carboplatin,-AUC=6, once every three weeks; 175 mg/m2 onceevery two weeks, plus 2000 mg/m² gemcitabine and 50 mg/m² epirubicin,once every two weeks; and 75 mg/m² weekly, three out of four weeks, pluscarboplatin, AUC=2, weekly, three out of four weeks.

In some embodiments, the nanoparticle composition of the taxane and thechemotherapeutic agent is administered according to any of the dosingregimes described in Table 1.

In some embodiments, there is provided a method of treating breastcancer in an individual comprising administering to the individual: a)an effective amount of a composition comprising nanoparticles comprisinga taxane (such as paclitaxel) and an albumin, and b) an effective amountof at least one other chemotherapeutic agent as provided in Rows 1 to 35in Table 1. In some embodiments, the administration of the nanoparticlecomposition and the chemotherapeutic agent may be any of the dosingregimes as indicated in Rows 1 to 35 in Table 1. In some embodiments,there is provided a method of treating metastatic breast cancer in anindividual comprising administering to the individual: a) an effectiveamount of a composition comprising nanoparticles comprising a taxane(such as paclitaxel) and an albumin, and b) an effective amount of atleast one other chemotherapeutic agent as provided in Rows 2, 4-8, and10-15 in Table 1. In some embodiments, the administration of thenanoparticle composition and the chemotherapeutic agent may be any ofthe dosing regimes as indicated in Rows 2, 4-8, and 10-15 in Table 1.

In some embodiments, there is provided a method of treating advancedbreast cancer in an individual comprising administering to theindividual: a) an effective amount of a composition comprisingnanoparticles comprising a taxane (such as paclitaxel) and an albumin,and b) an effective amount of at least one other chemotherapeutic agentas provided in Rows 1 and 16 in Table 1. In some embodiments, theadministration of the nanoparticle composition and the chemotherapeuticagent may be any of the dosing regimes as indicated in Rows 1 and 16 inTable 1. In some embodiments, there is provided a method of treatingstage IV breast cancer in an individual comprising administering to theindividual: a) an effective amount of a composition comprisingnanoparticles comprising a taxane (such as paclitaxel) and an albumin,and b) an effective amount of at least one other chemotherapeutic agentas provided in Row 3 in Table 1. In some embodiments, the administrationof the nanoparticle composition and the chemotherapeutic agent may bethe dosing regime as indicated in Row 3 in Table 1.

In some embodiments, there is provided a method of treating breastcancer in an individual in an adjuvant setting comprising administeringto the individual: a) an effective amount of a composition comprisingnanoparticles comprising a taxane (such as paclitaxel) and an albumin,and b) an effective amount of at least one other chemotherapeutic agentas provided in Rows 18 to 24 in Table 1. In some embodiments, theadministration of the nanoparticle composition and the chemotherapeuticagent may be any of the dosing regimes as indicated in Rows 18 to 24 inTable 1.

In some embodiments, there is provided a method of treating breastcancer in an individual in a neoadjuvant setting comprisingadministering to the individual: a) an effective amount of a compositioncomprising nanoparticles comprising a taxane (such as paclitaxel) and analbumin, and b) an effective amount of at least one otherchemotherapeutic agent as provided in Rows 25 to 35 in Table 1. In someembodiments, the administration of the nanoparticle composition and thechemotherapeutic agent may be any of the dosing regimes as indicated inRows 25 to 35 in Table 1.

In some embodiments, there is provided a method of treating lung cancerin an individual comprising administering to the individual: a) aneffective amount of a composition comprising nanoparticles comprising ataxane (such as paclitaxel) and an albumin, and b) an effective amountof at least one other chemotherapeutic agent as provided in Rows 36 to48 in Table 1. In some embodiments, the administration of thenanoparticle composition and the chemotherapeutic agent may be any ofthe dosing regimes as indicated in Rows 36 to 48 in Table 1.

In some embodiments, there is provided a method of treating NSCLC(including advanced NSCLC and first line NSCLC) in an individualcomprising administering to the individual: a) an effective amount of acomposition comprising nanoparticles comprising a taxane (such aspaclitaxel) and an albumin, and b) an effective amount of at least oneother chemotherapeutic agent as provided in Rows 36-40 and 42-43 inTable 1. In some embodiments, the administration of the nanoparticlecomposition and the chemotherapeutic agent may be any of the dosingregimes as indicated in Rows 36-40 and 42-43 in Table 1. In someembodiments, there is provided a method of treating advanced solid tumormalignancy in the lung in an individual comprising administering to theindividual: a) an effective amount of a composition comprisingnanoparticles comprising a taxane (such as paclitaxel) and an albumin,and b) an effective amount of at least one other chemotherapeutic agentas provided in Row 41 in Table 1. In some embodiments, theadministration of the nanoparticle composition and the chemotherapeuticagent may be the dosing regimes as indicated in Row 41 in Table 1. Insome embodiments, there is provided a method of treating SCLC in anindividual comprising administering to the individual: a) an effectiveamount of a composition comprising nanoparticles comprising a taxane(such as paclitaxel) and an albumin, and b) an effective amount of atleast one other chemotherapeutic agent as provided in Row 48 in Table 1.In some embodiments, the administration of the nanoparticle compositionand the chemotherapeutic agent may be the dosing regimes as indicated inRow 48 in Table 1.

In some embodiments, there is provided a method of treating ovariancancer in an individual comprising administering to the individual: a)an effective amount of a composition comprising nanoparticles comprisinga taxane (such as paclitaxel) and an albumin, and b) an effective amountof at least one other chemotherapeutic agent as provided in Rows 49 to52 in Table 1. In some embodiments, the administration of thenanoparticle composition and the chemotherapeutic agent may be any ofthe dosing regimes as indicated in Rows 49 to 52 in Table 1.

In some embodiments, there is provided a method of treating head andneck cancer in an individual comprising administering to the individual:a) an effective amount of a composition comprising nanoparticlescomprising a taxane (such as paclitaxel) and an albumin, and b) aneffective amount of at least one other chemotherapeutic agent asprovided in Rows 53 to 55 in Table 1. In some embodiments, theadministration of the nanoparticle composition and the chemotherapeuticagent may be any of the dosing regimes as indicated in Rows 53 to 55 inTable 1.

In some embodiments, there is provided a method of treating solid tumor(including advanced solid tumor) in an individual comprisingadministering to the individual: a) an effective amount of a compositioncomprising nanoparticles comprising a taxane (such as paclitaxel) and analbumin, and b) an effective amount of at least one otherchemotherapeutic agent as provided in Rows 56.to 59 in Table 1. In someembodiments, the administration of the nanoparticle composition and thechemotherapeutic agent may be any of the dosing regimes as indicated inRows 56 to 59 in Table 1.

In some embodiments, there is provided a method of treating melanoma(including metastatic melanoma) in an individual comprisingadministering to the individual: a) an effective amount of a compositioncomprising nanoparticles comprising a taxane (such as paclitaxel) and analbumin, and b) an effective amount of at least one otherchemotherapeutic agent as provided in. Rows 60-63 in Table 1. In someembodiments, the administration of the nanoparticle composition and thechemotherapeutic agent may be any of the dosing regimes as indicated inRows 60 to 63 in Table 1.

In some embodiments, there is provided a method of treating metastaticcolorectal cancer in an individual comprising administering to theindividual: a) an effective amount of a composition comprisingnanoparticles comprising a taxane (such as paclitaxel) and an albumin,and b) an effective amount of at least one other chemotherapeutic agentas provided in Row 64 in Table 1. In some embodiments, theadministration of the nanoparticle composition and the chemotherapeuticagent may be the dosing regime as indicated in Row 64 in Table 1.

In some embodiments, there is provided a method of treating pancreaticcancer in an individual comprising administering to the individual: a)an effective amount of a composition comprising nanoparticles comprisinga taxane (such as paclitaxel) and an albumin, and b) an effective amountof at least one other chemotherapeutic agent as provided in Rows 65 to66 in Table 1. In some embodiments, the administration of thenanoparticle composition and the chemotherapeutic agent may be any ofthe dosing regimes as indicated in Rows 65 to 66 in Table 1. TABLE 1 RowStudy therapy No. Combination Regime/Dosage type Protocol title 1. ABX +Carboplatin + Herceptin ® ABX: 100 mg/m² D1, 8, 15 Advanced A phase IIstudy of q4wk × 6 HER2+ Breast weekly dose-dense Carbo: AUC = 2 D1, 8,15 Cancer nanoparticle paclitaxel q4wk × 6 (ABI-007) Herceptin ®: 4mg/kg on wk 1, 2 mg/kg carboplatin ™, with all subsequent weeksHerceptin ® as first or second-line therapy of advanced HER2+ breastcancer 2. ABX alone ABX: 125 mg/m² Metastatic Phase II trial of weekly(+Herceptin ®) qwk × ¾ Breast Cancer Abraxane ™ monotherapy for 1st-line MBC (plus Herceptin ® in HER2+ pts) 3. ABX + Navelbine ® L1: ABX:80 mg/m Stage IV Phase I-II study weekly (±G-CSF) Nav: 15 mg/m² BreastCancer ABX + Navelbine ®, L2: ABX: 90 mg/m² with or without G-CSF, Nav:20 mg/m² in stage IV breast L3: ABX: 100 mg/m² cancer Nav: 22.5 mg/m²L4: ABX: 110 mg/m² Nav: 25 mg/m² L5: ABX: 125 mg/m² Nav: 25 mg/m² qwkall levels 4. ABX + Xeloda ® ABX: 125 mg/m² qwk × ⅔ Metastatic Phase II1st-line ABX + Xeloda ® Xeloda ®: 825 mg/m² D1-14 Breast Cancer MBCtrial q3wk 5. ABX + Anthracycline Metastatic Phase I/II trial ABX BreastCancer plus Doxil ® for MBC plus limited PK 6. ABX + Gemcitabine ABX:125 mg/m² Metastatic Randomized Phase II Gem: 1000 mg/m2 Breast CancerTrial of Weekly nab qwk × ⅔ (nanoparticle albumin bound)-Paclitaxel(nab- paclitaxel) in Combination with Gemcitabine in Patients with HER2Negative Metastatic Breast Cancer 7. ABX + Lapatinib Metastatic PhaseI/II Abraxane ™ + GW572016 Breast Cancer 8. ABX + Lapatinib ABX: 100mg/m² qwk × ¾ Metastatic Phase I dose escalation Lapatinib: starting at1000 mg/d × 2 Breast Cancer study of a 2 day oral days lapatinibchemosensitization pulse given prior to weekly intravenous Abraxane ™ inpatients with advanced solid tumors 9. ABX + FEC ABX: 220 mg/m² q2wk × 6Breast Cancer Phase II preoperative (+Herceptin ®) followed by trial ofAbraxane ™ FEC: 4 cycles (+Herceptin ® for followed by FEC HER2+ pts)(+Herceptin ® as appropriate) in breast cancer 10. ABX + Carboplatin +Avastin ® ABX: 100 mg/m² qwk D1, 8, 15 Metastatic Phase II safety andCarbo: AUC = 2 qwk D1, 8, 15 Breast Cancer tolerability study ofAvastin ®: 10 mg/m² q2wk (HER2-, ER-, Abraxane ™, Avastin ® PR-) andcarboplatin in triple negative metastatic breast cancer patients 11.ABX + Avastin ® ABX: 130 mg/m² qwk + Avastin ® Metastatic Three armphase II trial vs Breast Cancer in 1^(st) line HER2- ABX: 260 mg/m²q2wk + Avastin ® negative MBC patients vs ABX: 260 mg/m² q3wk +Avastin ® 12. ABX + Avastin ® ABX: 125 mg/m² qwk × ¾ + Avastin ®Metastatic Single arm study of Breast Cancer Abraxane ™ and Avastin ® in1^(st) line MBS 13. ABX + Avastin ® ABX + Avastin ® qwk MetastaticRandomized Phase III vs Breast Cancer trial in 1^(st) line and 2^(nd)Taxol ® + Avastin ® qwk line MBC with biological correlates analysis 14.ABX + Xeloda ® + Lapatinib Metastatic Phase II Abraxane ™ in BreastCancer combination with Xeloda ® and Lapatinib for metastatic breastcancer 15. ABX + Gemcitabine ABX: 3000 mg/m² D1 q3wk Metastatic Singlearm Phase II Gem: 1250 mg/m² D1, 8 q3wk Breast Cancer study ofAbraxane ™ and gemcitabine for 1^(st) line MBC 16. ABX + RAD001 AdvancedPhase I/II study of Breast Cancer Abraxane ™ in combination with RAD001in patients with advanced breast cancer 17. ABX + Sutent ® Breast CancerPhase I study of Abraxane ™ in combination with Sutent ® 18. ABX + AC +G- AC + G-CSF q2wk × 4 Breast Cancer - Abraxane ™ in dose- CSF(+Herceptin ®) followed by Adjuvant dense adjuvant ABX: 260 mg/m² q2wk ×4 chemotherapy for early (+Herceptin ® for HER2+ pts) stage breastcancer 19. ABX + AC + G- Dose dense AC + G-CSF Breast Cancer - Phase IIpilot adjuvant CSF (+Herceptin ®) followed by ABX Adjuvant trial ofAbraxane ™ in (+Herceptin ® for HER2+ pts) breast cancer qwk 20. ABX +AC AC followed by ABX: 260 mg/m² Breast Cancer - Adjuvant Dose dense vsAdjuvant Registrational Trial AC followed by Taxol ® Rx length 16 wks21. ABX + AC AC q2wk followed by Breast Cancer - Phase II dose dense(+G-CSF) ABX: 260 mg/m² + G-CSF Adjuvant pilot adjuvant study of q2wkAbraxane ™ in breast Rx length 16 wks cancer 22. ABX + AC Dose dense ACfollowed by Breast Cancer - Pilot adjuvant breast (+Avastin ®) ABX(+Avastin ® in HER2+ Adjuvant cancer study pts) 23. ABX + AC AC BreastCancer - BIG study: Dose dense followed by ABX Adjuvant vs standardadjuvant q2wk or q3wk chemotherapy 24. ABX (ABI-007) + AC + Neulasta ®AC followed by Breast Cancer - Phase II - Pilot Study ABX q2wk × 4Adjuvant Evaluating the Safety of a Dose-Dense Regime - AC × 4 =>ABI-007 × 4 Q 2 WEEKS + Neulasta ® - Given as Adjuvant Chemotherapy ofHigh- Risk Women with Early Breast Cancer 25. ABX + FEC ABX: 100 mg/m²qwk × 12 Locally A Phase II Study of (+Herceptin ®) followed by AdvancedBreast Neoadjuvant 5-FU: 500 mg/m² q3wk Cancer - Chemotherapy withEpirubicin: 100 mg/m² Neoadjuvant Sequential Weekly (withoutHerceptin ®) Nanoparticle Albumin or Bound Paclitaxel Epirubicin: 75mg/m² (Abraxane ™) Followed (with Herceptin ® for HER2+ by5-Fluorouracil, pts) Epirubicin, Cyclophosphamide: 500 mg/m²Cyclophosphamide q3wk (FEC) in Locally Advanced Breast Cancer 26. ABX +Gemcitabine + Epirubicin Arm 1: Neoadjuvant: Gem: 2000 mg/m², BreastCancer - Phase II Trial of Dose ABX: 175 mg/m², Epi Neoadjuvant DenseNeoadjuvant 50 mg/m² Gemcitabine, q2wk × 6 Epirubicin, ABI-007 Arm 2:Adjuvant: Gem: 2000 mg/m², (GEA) in Locally ABX: 220 mg/m² Advanced orq2wk × 4 Inflammatory Breast Cance 27. ABX + Herceptin ® ABX: 260 mg/m²q2wk + Herceptin ® Breast Cancer - Phase II Multi-center followed byNeoadjuvant study neoadjuvant. Navelbine ® + Herceptin ® 28. ABX +Carboplatin TAC Breast Cancer - 3 arms Randomized (+Herceptin ®) + AC vsNeoadjuvant dose dense phase II AC followed by ABX + carbo trial ofneoadjuvant vs chemotherapy in AC followed by ABX + carbo + Herceptin ®patients with breast cancer 29. ABX + Capecitabine ABX: 260 mg/m² q3wk ×4 Breast Cancer - Phase II neoadjuvant Xeloda ® 850 mg/m² D1-14Neoadjuvant trial of Abraxane ™ and q3wk × 4 capecitabine in locallyadvanced breast cancer 30. ABX + Carboplatin ABX qwk Breast Cancer -Phase I/II trial of (+Avastin ®) carbo qwk + Neoadjuvant neoadjuvantAvastin ® in HER2+ pts chemotherapy (NCT) with weekly nanoparticlepaclitaxel (ABI-007, Abraxane ™) in combination with carboplatin andAvastin ® in clinical stage I-III. 31. ABX + Carboplatin + Herceptin ® +ABX: 100 mg/m² qwk × ¾ Breast Cancer - Phase II study of Avastin ®Carbo: AUC = 5 + Herceptin ® + Avastin ® Neoadjuvant weekly bevacizumab4 week cycle × 6 administered with weekly trastuzumab, ABI-007, andcarboplatin as preoperative therapy in HER2-neu gene amplified breastcancer tumors 32. ABX + Lapatinib ABX: 260 mg/m² q3wk Breast Cancer -Pilot neoadjuvant trial Lapatinib: 1000 mg/day Neoadjuvant withcombination of ABI-007 (Abraxane ™) and GW572016 (Lapatinib) 33. ABX +Capecitabine ABX: 200 mg/m² Breast Cancer - Phase II neoadjuvant q3wk ×4 Neoadjuvant trial of Abraxane ™ and Xeloda ®: 1000 mg/m² capecitabinein locally D1-14 q3wk × 4 advanced breast cancer 34. ABX ± Avastin ® +AC ABX qwk ± Avastin ® followed Breast Cancer - Phase III trial of(+G-CSF) by A qwk + C daily Neoadjuvant paclitaxel vs vs Abraxane ™ withor Taxol ® qwk ± Avastin ® without Avastin ® in followed by A qwk + Cdaily combination with doxorubicin and cyclophosphamide plus G-CSF 35.ABX + AC ABX followed by AC Breast Cancer - Phase II neoadjuvantNeoadjuvant trial with gene expression analyses 36. ABX + Carboplatin +Avastin ® ABX: 300 mg/m² q3wk 1^(st) line An open label phase II Carbo:AUC = 6 q3wk Advanced trial of Abraxane ™, Avastin ®: 15 mg/kg NSCLCcarboplatin and 4 cycles Avastin ® in patients with advanced non-squamous non-small cell lung cancer 37. ABX + Carboplatin L1: ABX: 225mg/m² Advanced Phase II toxicity pilot L2: ABX: 260 mg/m² NSCLC study ofAbraxane ™ L3: ABX: 300 mg/m² and carboplatin in Cohorts 1-4: ABX q3wkadvanced non-small cell Cohorts 5-7: ABX weekly lung cancer. Cohort 8:75 additional patients Carbo fixed at AUC = 6 q3wk 38. ABX + CarboplatinCarbo: AUC = 6 + ABX 1^(st) line NSCLC Phase III Registration - vs NSCLC1^(st) line therapy Carbo: AUC = 6 + Taxol ®: 225 mg/m² 39. ABX +Carboplatin ABX: 100 mg/m² d1, 8, 15 1^(st) line NSCLC Phase II Trial ofweekly Carbo: AUC = 6 q4wk Abraxane ™ plus Amendment: ABX: 125 mg/m²,carboplatin in 1st-line D1, 8, 15 NSCLC 40. ABX + Carboplatin +Avastin ® Weekly NSCLC 41. ABX + Carboplatin Arm 1: ABX: 100, 125, 150mg/m² Lung Cancer - Phase I Trial of D1, 8, 15 q4wk Advanced Solidcarboplatin and Arm 2: ABX 220, 260, 300, 340 mg/m² Tumor Abraxane ™ ona q3wk Malignancy weekly and every three Arm 3: ABX 100, 125, 150 mg/m²week schedule in D1, 8 patients with Advanced Carbo: AUC = 6 in all armsSolid Tumor Malignancies 42. ABX + Gemcitabine NSCLC Abraxane ™ in orcombination with ABX + Avastin ® gemcitabine or Avastin ® 43. ABX +Gemcitabine NSCLC Phase I trial of Abraxane ™ in combination withgemcitabine 44. ABX + Carboplatin + Avastin ® ABX: 225, 260, 300 mg/m²Lung Cancer Phase I/II study of Carbo: AUC = 6 Abraxane ™ and q3wk +Avastin ® carboplatin AUC 6, plus Avastin ® (Standard 3 + 3 Phase Idesign; PhII: 40 pts) 45. ABX + Alimta ® ABX: 220, 260, 300 mg/m² LungCancer Phase I/II study of q3wk Abraxane ™ + Alimta ® Pemtrexed: 500 mgq3wk for 2nd-line NSCLC 46. ABX + Cisplatin Lung Cancer Phase I/II trialof Abraxane ™ plus cisplatin in advanced NSCLC 47. ABX + Navelbine ® +Cisplatin Lung Cancer Phase I/II study of Abraxane ™, Navelbine ®, andCisplatin for treatment of advanced NSCLC 48. ABX + Carboplatin ABX: 300mg/m² q3wk SCLC Phase II trial of Carbo: AUC = 6 q3wk Abraxane ™ andcarboplatin in extensive stage small cell lung cancer 49. ABX +Carboplatin ABX: 100 mg/m² qwk × ¾ Ovarian Cancer A phase II trial ofCarbo: AUC = 6 Abraxane ™ + Carboplatin in recurrent ovarian cancer 50.ABX + Carboplatin ABX: qwk Ovarian Cancer Phase I study of ABX: q3wAbraxane ™ plus carbo for treatment of advanced ovarian Carbo: AUC = 6both arms cancer 51. ABX + Carboplatin ABX: TBD by ABI-CA034 OvarianCancer 1st line, optimally vs debulked, registration Taxol ® 175 mg/m²trial. Carbo AUC 6 + ABX Carbo: AUC = 6 in both arms vs Carbo + Taxol ®175 mg/m². Endpoint: relapse free survival, survival 52. ABX + Avastin ®ABX: 100 mg/m² qwk × ¾ Ovarian Cancer Phase II study of Avastin ®: 10mg/m² q2wk bevacizumab with Abraxane ™ in patients with recurrent,platinum resistant primary epithelial ovarian or primary peritonealcarcinoma 53. ABX + 5-FU + Cisplatin ABX: D1 Head and Neck Unresectablelocalized 5-FU: 750 mg/m² CIV × 5 Cancer head and neck cancer cisplatin:75 mg/m² D1 Phase II Abraxane ™ in followed by XRT/surgery combinationwith 5-FU and cisplatin 54. ABX + 5-FU + Cisplatin 5-FU: 750 mg/m² CIV ×5 Head and Neck Unresectable localized cisplatin: 75 mg/m² D1 ± ABXCancer head and neck cancer D1 Phase III 5-FU and followed byXRT/surgery cisplatin with or without Abraxane ™ 55. ABX + CetuximabHead and Neck Phase II multicenter trial Cancer of Abraxane ™ incombination with cetuximab in 1^(st) line treatment of locally advancedor metastatic head and neck cancer 56. ABX + Rapamycin ABX: 100 mg/m²qwk Solid Tumors Phase I Study of Rapamycin: 5-40 mg dose Rapamycin inescalation Combination with Abraxane ™ in Advanced Solid Tumors 57.ABX + Satraplatin Solid Tumors Phase I trial of Abraxane ™ andSatraplatin 58. ABX + Gemcitabine ABX: 180, 220, 260, 300, 340 mg/m²Advanced Solid Phase I Trial of q3wk Tumors Abraxane ™ in Gemcitabine:1000 mg/m² D1 combination with and D8 Gemcitabine 59. ABX + GefitinibABX: 100 mg/m² qwk × ¾ Advanced Solid Phase I dose escalation Gefitinibstarting at 1000 mg/d × 2 Tumors study of gefitinib chemosensitizationpulse given prior to weekly Abraxane ™ 60. ABX + Avastin ® MetastaticPhase II study of Melanoma Abraxane ™ and Avastin ® in metastaticmelanoma 61. ABX + Avastin ® Melanoma Abraxane ™ and Avastin ® astherapy for patients with malignant melanoma 62. ABX + CarboplatinMetastatic Phase II study of Melanoma Abraxane ™ and carboplatin inmetastatic melanoma 63. ABX + Sorafenib + Carboplatin ABX: qwkMetastatic Phase II study of Sorafenib: D2-19 Melanoma Abraxane ™ inCarbo: AUC = 6 D1 combination with carboplatin and sorafenib inmetastatic melanoma 64. ABX + Capecitabine Metastatic Phase II trial ofColorectal Abraxane ™ in Cancer (after combination with failure ofXeloda ® for previously oxaliplatin- treated patient with based therapyadvance or metastatic and irinotecan- colorectal cancer based therapy)65. ABX + Gemcitabine Weekly Pancreatic Phase I study of CancerAbraxane ™ in combination with gemcitabine in pancreatic cancer 66.ABX + Gemcitabine ABX + Gem Pancreatic Phase III registration trial vsCancer in pancreatic cancer Gem 67. ABX + anti- Abraxane ™ combinedangiogenic with anti-angiogenic agents agents, e.g. Avastin ® 68. ABX +proteasome Abraxane ™ combined inhibitors with proteasome inhibitors,e.g. Velcade ® 69. ABX + EGFR Abraxane ™ combined inhibitors with EGFRinhibitors, e.g. Tarceva ®

As used in herein (for example in Table 1), ABX refers to Abraxane™;GW572016 refers to lapatinib; Xel refers to capecitabine or Xeloda®;bevacizumab is also known as Avastin®; trastuzumab is also known asHerceptin®; pemtrexed is also known as Alimta®; cetuximab is also knownas Erbitux®; gefitinib is also known as Iressa®; FEC refers to acombination of 5-fluorouracil, Epirubicin and Cyclophosphamide; ACrefers to a combination of Adriamycin plus Cyclophosphamide; TAC refersto a FDA approved adjuvant breast cancer regime; RAD001 refers to aderivative of rapamycin; NSCLC refers to non-small cell lung cancer; andSCLC refers to small cell lung cancer.

As used herein (for example in Table 1), AUC refers to area under curve;q4 wk refers to a dose every 4 weeks; q3 wk refers to a dose every 3weeks; q2 wk refers to a dose every 2 weeks; qwk refers to a weeklydose; qwk×3/4 refers to a weekly dose for 3 weeks with the 4^(th) weekoff; qwk×2/3 refers to a weekly dose for 2 weeks with the 3^(rd) weekoff.

Combination Therapy with Radiation Therapy and Surgery

In another aspect, the present invention provides a method of treatingproliferative disease (such as cancer) comprising a first therapycomprising administering a taxane (particularly nanoparticles comprisinga taxane) and a carrier protein and a second therapy comprisingradiation and/or surgery.

In some embodiments, the method comprises: a) a first therapy comprisingadministering to the individual a composition comprising nanoparticlescomprising an effective amount of a taxane and a carrier protein (suchas albumin) and b) a second therapy comprising radiation therapy,surgery, or combinations thereof. In some embodiments, the taxane iscoated with the carrier protein (such as albumin). In some embodiments,the second therapy is radiation therapy. In some embodiments, the secondtherapy is surgery.

In some embodiments, the method comprises a) a first therapy comprisingadministering to the individual a composition comprising nanoparticlescomprising paclitaxel and an albumin; and b) a second therapy comprisingradiation therapy, surgery, or combinations thereof. In someembodiments, the second therapy is radiation therapy. In someembodiments, the second therapy is surgery. In some embodiments, thepaclitaxel/albumin nanoparticles have an average diameter of no greaterthan about 200 nm. In some embodiments, the paclitaxel/albuminnanoparticle composition is substantially free (such as free) ofsurfactant (such as Cremophor). In some embodiments, the weight ratio ofthe albumin to paclitaxel in the composition is about 18:1 or less, suchas about 9:1 or less. In some embodiments, the paclitaxel is coated withalbumin. In some embodiments, the paclitaxel/albumin nanoparticles havean average diameter of no greater than about 200 nm and thepaclitaxel/albumin composition is substantially free (such as free) ofsurfactant (such as Cremophor). In some embodiments, thepaclitaxel/albumin nanoparticles have an average diameter of no greaterthan about 200 nm and the paclitaxel is coated with albumin. In someembodiments, the nanoparticle composition is Abraxane™.

The administration of the nanoparticle composition may be prior to theradiation and/or surgery, after the radiation and/or surgery, orconcurrent with the radiation and/or surgery. For example, theadministration of the nanoparticle composition may precede or follow theradiation and/or surgery therapy by intervals ranging from minutes toweeks. In some embodiments, the time period between the first and thesecond therapy is such that the taxane and the radiation/surgery wouldstill be able to exert an advantageously combined effect on the cell.For example, the taxane (such as paclitaxel) in the nanoparticlecomposition may be administered less than about any of 1, 3, 6, 9, 12,18, 24, 48, 60, 72, 84, 96, 108, 120 hours prior to the radiation and/orsurgery. In some embodiments, the nanoparticle composition isadministered less than about 9 hours prior to the radiation and/surgery.In some embodiments, the nanoparticle composition is administered lessthan about any of 1, 2, 3, 4, 5, 6, 7, 8, 9, or 10 days prior to theradiation/surgery. In some embodiments, the taxane (such as paclitaxel)in the nanoparticle composition is administered less than about any of1, 3, 6, 9, 12, 18, 24, 48, 60, 72, 84, 96, 108, or 120 hours after theradiation and/or surgery. In some embodiments, it may be desirable toextend the time period for treatment significantly, where several daysto several weeks lapse between the two therapies.

Radiation contemplated herein includes, for example, γ-rays, X-rays(external beam), and the directed delivery of radioisotopes to tumorcells. Other forms of DNA damaging factors are also contemplated such asmicrowaves and UV irradiation are also contemplated. Radiation may begiven in a single dose or in a series of small doses in adose-fractionated schedule. The amount of radiation contemplated hereinranges from about 1 to about 100 Gy, including, for example, about 5 toabout 80, about 10 to about 50 Gy, or about 10 Gy. The total dose may beapplied in a fractioned regime. For example, the regime may comprisefractionated individual doses of 2 Gy. Dosage ranges for radioisotopesvary widely, and depends on the half-life of the isotope and thestrength and type of radiation emitted.

When the radiation comprises use of radioactive isotopes, the isotopemay be conjugated to a targeting agent, such as a therapeutic antibody,which carries the radionucleotide to the target tissue. Suitableradioactive isotopes include, but are not limited to, astatine²¹¹,¹⁴carbon, ⁵¹chromium, ³⁶chlorine, ⁵⁷iron, ⁵⁸cobalt, copper⁶⁷, ¹⁵²Eu,gallium⁶⁷, ³hydrogen, iodine¹²³, iodine¹³¹, indium¹¹¹, ⁵⁹ion,³²phosphorus, rhenium¹⁸⁶, ⁷⁵selenium, ³⁵sulphur, technicium^(99m),and/or yttrium⁹⁰.

In some embodiments, enough radiation is applied to the individual so asto allow reduction of the normal dose of the taxane (such as paclitaxel)in the nanoparticle composition required to effect the same degree oftreatment by at least about any of 5%, 10%, 20%, 30%, 50%, 60%, 70%,80%, 90%, or more. In some embodiments, enough taxane in thenanoparticle composition is administered so as to allow reduction of thenormal dose of the radiation required to effect the same degree oftreatment by at least about any of 5%, 10%, 20%, 30%, 50%, 60%, 70%,80%, 90%, or more. In some embodiments, the dose of both the taxane(such as paclitaxel) in the nanoparticle composition and the radiationare reduced as compared to the corresponding normal dose of each whenused alone.

In some embodiments, the combination of administration of thenanoparticle composition and the radiation therapy producesupra-additive effect. In some embodiments, the taxane (such aspaclitaxel) in the nanoparticle composition is administered once at thedose of 90 mg/kg, and the radiation is applied five times at 80 Gydaily.

Surgery described herein includes resection in which all or part ofcancerous tissue is physically removed, exercised, and/or destroyed.Tumor resection refers to physical removal of at least part of a tumor.In addition to tumor resection, treatment by surgery includes lasersurgery, cryosurgery, electrosurgery, and micropically controlledsurgery (Mohs surgery). Removal of superficial surgery, precancers, ornormal tissues are also contemplated.

The radiation therapy and/or surgery may be carried out in addition tothe administration of chemotherapeutic agents. For example, theindividual may first be administered with a taxane-containingnanoparticle composition and at least one other chemotherapeutic agent,and subsequently be subject to radiation therapy and/or surgery.Alternatively, the individual may first be treated with radiationtherapy and/or surgery, which is then followed by the administration ofa nanoparticle composition and at least one other chemotherapeuticagent. Other combinations are also contemplated.

Administration of nanoparticle compositions disclosed above inconjunction with administration of chemotherapeutic agent is equallyapplicable to those in conjunction with radiation therapy and/orsurgery.

In some embodiments, the nanoparticle composition of the taxane and/orthe chemotherapeutic agent is administered in conjunction with radiationaccording to any of the dosing regimes described in Table 2.

In some embodiments, there is provided a method of treating NSCLC in anindividual comprises a) a first therapy comprising administering to theindividual a composition comprising nanoparticles comprising taxane(such as paclitaxel) and an albumin; and b) a second therapy comprisingradiation as provided in Rows 1 to 5 in Table 2. In some embodiments,the administration of the nanoparticle composition and thechemotherapeutic agent may be any of the dosing regimes as indicated inRows 1 to 5 in Table 2.

In some embodiments, there is provided a method of treating head andneck cancer in an individual comprises a) a first therapy comprisingadministering to the individual a composition comprising nanoparticlescomprising taxane (such as paclitaxel) and an albumin; and b) a secondtherapy comprising radiation as provided in Rows 6 to 9 in Table 2. Insome embodiments, the administration of the nanoparticle composition andthe chemotherapeutic agent may be any of the dosing regimes as indicatedin Rows 6 to 9 in Table 2.

In some embodiments, there is provided a method of treating pancreaticcancer in an individual comprises a) a first therapy comprisingadministering to the individual a composition comprising nanoparticlescomprising taxane (such as paclitaxel) and an albumin; and b) a secondtherapy comprising radiation as provided in Row 10 in Table 2. In someembodiments, the administration of the nanoparticle composition and thechemotherapeutic agent may be the dosing regimes as indicated in Row 10in Table 2.

In some embodiments, there is provided a method of treating gastricmalignancies in an individual comprises a) a first therapy comprisingadministering to the individual a composition comprising nanoparticlescomprising taxane (such as paclitaxel) and an albumin; and b) a secondtherapy comprising radiation as provided in Row 11 in Table 2. In someembodiments, the administration of the nanoparticle composition and thechemotherapeutic agent may be the dosing regimes as indicated in Row 11in Table 2. TABLE 2 Row Study therapy No. Combination Regime/Dosage typeProtocol title 1 ABX + Radiation NSCLC Phase I/II trial of Abraxane ™combined with radiation 2 ABX + Carboplatin + Radiation NSCLC Phase I/IItrial of Abraxane ™ and carboplatin combined with radiation. 3 ABX +Carboplatin + Radiation 1 cycle ABX/Carbo induction NSCLC Phase IIchemoradiation followed by in NSCLC 2 or 3 times weekly pulse ABX +radiation 4 ABX + Carboplatin + Radiation NSCLC Abraxane ™/ carboplatininduction followed by Abraxane ™ + radiation in stage III A&B PS2 NSCLCpatients 5 ABX + Carboplatin + Radiation ABX qwk + carbo + radiationNSCLC Phase II study followed by ABX q3wk + carbo 6 ABX + Radiation Headand Abraxane ™ as a Neck Cancer radiosensitizer in head and neck cancer7 ABX + Cetuximab + Radiation Head and PhaseI/II Abraxane ™ Neck Cancerin combination with cetuximab and radiation 8 ABX + Carboplatin + 5-Induction: ABX 135 mg/m² Head and Phase I/II study of FU + Hydroxyurea +Radiation qwk + carbo: AUC = 2 Neck Cancer induction chemotherapyfollowed by with Abraxane ™ and Concurrent chemoradiation: carboplatinfollowed by ABX: 100 mg/m² concomitant 5-FU: 600 mg/m² fluorouracil,hydroxyurea: 5000 mg BID hydroxyurea, Abraxane ™ and IMRT for locallyadvanced head and neck cancers 9 ABX + Carboplatin + Erbitux ® + ABX:20-50 mg/m² qwk × 7 Locally Phase I trial of Radiation dose escalationAdvanced Abraxane ™ in Eribitux ®: 400 mg/m² day 7, Head and combinationwith 250 mg/m² qwk × 7 Neck Cancer carboplatin, cetuximab Carbo: AUC =1.5 qwk × 7 and IMRT in locally IMRT advanced squamous cell cancer ofthe head and neck 10 ABX + Gemcitabine + Radiation qwk Pancreatic Arandomized phase II Cancer trial of weekly gemcitabine, Abraxane ™, andexternal irradiation for locally advanced pancreatic cancer 11 ABX +Cisplatin + Radiation Gastric Phase I/II combination Malignancies ofAbraxane ™/cisplatin and radiation for patients with resectedgastric/GEJ malignancies.

In some embodiments, the invention provides pharmaceutical compositionscomprising nanoparticles comprising a taxane (such as paclitaxel) and acarrier protein (such as albumin) for use in the treatment of aproliferative disease (such as cancer), wherein said use comprises asecond therapy comprising radiation therapy, surgery, or combinationsthereof.

Metronomic Therapy

The invention also provides metronomic therapy regime. There is provideda method of administering to an individual a composition comprisingnanoparticles comprising a taxane (such as paclitaxel, docetaxel, orortataxel) and a carrier protein (such as albumin) based on a metronomicdosing regime. The methods are applicable to methods of treatment,delaying development, and other clinical settings and configurationsdescribed herein. For example, in some embodiments, the methods areuseful for treatment of proliferative diseases (such as cancer).

“Metronomic dosing regime” used herein refers to frequent administrationof a taxane at without prolonged breaks at a dose below the establishedmaximum tolerated dose via a traditional schedule with breaks(hereinafter also referred to as a “standard MTD schedule” or a“standard MTD regime”). In metronomic dosing, the same, lower, or highercumulative dose over a certain time period as would be administered viaa standard MTD schedule may ultimately be administered. In some cases,this is achieved by extending the time frame and/or frequency duringwhich the dosing regime is conducted while decreasing the amountadministered at each dose. Generally, the taxane administered via themetronomic dosing regime of the present invention is better tolerated bythe individual. Metronomic dosing can also be referred to as maintenancedosing or chronic dosing.

In some embodiments, there is provided a method of administering acomposition comprising nanoparticles comprising a taxane and a carrierprotein (such as albumin), wherein the nanoparticle composition isadministered over a period of at least one month, wherein the intervalbetween each administration is no more than about a week, and whereinthe dose of the taxane at each administration is about 0.25% to about25% of its maximum tolerated dose following a traditional dosing regime.In some embodiments, there is provided a method of administering acomposition comprising nanoparticles comprising paclitaxel and analbumin, wherein the nanoparticle composition is administered over aperiod of at least one month, wherein the interval between eachadministration is no more than about a week, and wherein the dose of thetaxane at each administration is about 0.25% to about 25% of its maximumtolerated dose following a traditional dosing regime.

In some embodiments, the dosing of the taxane (such as paclitaxel) inthe nanoparticle composition per administration is less than about anyof 1%, 2%, 3&, 4%, 5%, 6%, 7%, 8%, 9%, 10%, 11%, 12%, 13%, 14%, 15%,18%, 20%, 22%, 24%, 25%, of the MTD for the same taxane (such aspaclitaxel) in the same formulation following a given traditional dosingschedule. Traditional dosing schedule refers to the dosing schedule thatis generally established in a clinical setting. For example, thetradition dosing schedule for Abraxane™ is a three-weekly schedule,i.e., administering the composition every three weeks.

In some embodiments, the dosing of the taxane (such as paclitaxel) peradministration is between about 0.25% to about 25% of the correspondingMTD value, including for example any of about 0.25% to about 20%, about0.25% to about 15%, about 0.25% to about 10%, about 0.25% to about 20%,and about 0.25% to about 25%, of the corresponding MTD value. The MTDvalue for a taxane following a traditional dosing schedule is known orcan be easily determined by a person skilled in the art. For example,the MTD value when Abraxane™ is administered following a traditionalthree-week dosing schedule is about 300 mg/m ².

In some embodiments, there is provided a method of administering acomposition comprising nanoparticles comprising a taxane and a. carrierprotein (such as albumin), wherein the nanoparticle composition isadministered over a period of at least one month, wherein the intervalbetween each administration is no more than about a week, and whereinthe dose of the taxane at each administration is about 0.25 mg/m² toabout 25 mg/m². In some embodiments, there is provided a method ofadministering a composition comprising nanoparticles comprisingpaclitaxel and an albumin, wherein the nanoparticle composition isadministered over a period of at least one month, wherein the intervalbetween each administration is no more than about a week, and whereinthe dose of the taxane at each administration is about 0.25 mg/m² toabout 25 mg/m².

In some embodiments, the dose of the taxane (such as paclitaxel) at eachadministration is less than about any of 2, 3, 4, 5, 6, 7, 8, 9, 10, 11,12, 13, 14, 15, 18, 20, 22, 25, and 30 mg/m². For example, the dose ofthe taxane (such as paclitaxel) can range from about 0.25 mg/m² to about30 mg/m², about 0;25 mg/m² to about 25 mg/m², about 0.25 mg/m² to about15 mg/m², about 0.25 mg/m² to about 10 mg/m², and about 0.25 mg/m² toabout 5 mg/m².

Dosing frequency for the taxane (such as paclitaxel) in the nanoparticlecomposition includes, but is not limited to, at least about any of oncea week, twice a week, three times a week, four times a week, five timesa week, six times a week, or daily. Typically, the interval between eachadministration is less than about a week, such as less than about any of6, 5, 4, 3, 2, or 1 day. In some embodiments, the interval between eachadministration is constant. For example, the administration can becarried out daily, every two days, every three days, every four days,every five days, or weekly. In some embodiments, the administration canbe carried out twice daily, three times daily, or more frequent.

The metronomic dosing regimes described herein can be extended over anextended period of time, such as from about a month up to about threeyears. For example, the dosing regime can be extended over a period ofany of about 2, 3, 4, 5, 6, 7, 8, 9, 10, 11, 12, 18, 24, 30, and 36months. Generally, there are no breaks in the dosing schedule.

The cumulative dose of the taxane (such as paclitaxel) administered bythe metronomic regime may be higher than that of the taxane administeredaccording to a standard MTD dosing schedule over the same time period.In some embodiments, the cumulative dose of the taxane administered bythe metronomic regime equals to or is lower than that of the taxaneadministered according to a standard MTD dosing schedule over the sametime period.

It is understood that the teaching provided herein is for examples only,and that metronomic dosing regime can be routinely designed inaccordance with the teachings provided herein and based upon theindividual standard MTD schedule, and that the metronomic dosing regimeused in these experiments merely serves as one example of possiblechanges in dosing interval and duration which are made to a standard MTDschedule to arrive at an optimal metronomic dosing regime.

The metronomic dosing regime described herein may be used alone as atreatment of a proliferative disease, or carried out in a combinationtherapy context, such as the combination therapies described herein. Insome embodiments, the metronomic therapy dosing regime may be used incombination or conjunction with other established therapies administeredvia standard MTD regimes. By “combination or in conjunction with” it ismeant that the metronomic dosing regime of the present invention isconducted either at the same time as the standard MTD regime ofestablished therapies, or between courses of induction therapy tosustain the benefit accrued to the individual by the induction therapy,the intent is to continue to inhibit tumor growth while not undulycompromising the individual's health or the individual's ability towithstand the next course of induction therapy. For example, ametronomic dosing regime may be adopted after an initial short course ofMTD chemotherapy.

The nanoparticle compositions administered based on the metronomicdosing regime described herein can be administered to an individual(such as human) via various routes, such as parenterally, includingintravenous, intra-arterial, intrapulmonary, oral, inhalation,intravesicular, intramuscular, intra-tracheal, subcutaneous,intraocular, intrathecal, or transdermal. For example, the nanoparticlecomposition can be administered by inhalation to treat conditions of therespiratory tract. The composition can be used to treat respiratoryconditions such as pulmonary fibrosis, broncheolitis obliterans, lungcancer, bronchoalveolar carcinoma, and the like. In some embodiments,the nanoparticle composition is administered orally.

Some various exemplary embodiments are provided below.

In some embodiments, there is provided a method of administering acomposition comprising nanoparticles comprising a taxane and a carrierprotein (such as albumin), wherein the nanoparticle composition isadministered over a period of at least one month, wherein the intervalbetween each administration is no more than about a week, and whereinthe dose of the taxane at each administration is about 0.25% to about25% of its maximum tolerated dose following a traditional dosing regime.In some embodiments, the taxane is coated with the carrier protein (suchas albumin). In some embodiments, the dose of the taxane peradministration is less than about any of 1%, 2%, 3%, 4%, 5%, 6%, 7%, 8%,9%, 10%, 11%, 12%, 13%, 14%, 15%, 18%, 20%, 22%, 24%, or 25% of themaximum tolerated dose. In some embodiments, the taxane is administeredat least about any of 1×, 2×, 3×, 4×, 5', 6×, 7× (i.e., daily) a week.In some embodiments, the intervals between each administration are lessthan about any of 7 days, 6 days, 5 days, 4 days, 3 days, 2 days, and 1day. In some embodiments, the taxane is administered over a period of atleast about any of 2, 3, 4, 5, 6, 7, 8, 9, 10, 11, 12, 18, 24, 30 and 36months.

In some embodiments, there is provided a method of administering acomposition comprising nanoparticles comprising paclitaxel and analbumin, wherein the nanoparticle composition is administered over aperiod of at least one month, wherein the interval between eachadministration is no more than about a week, and wherein the dose of thetaxane at each administration is about 0.25% to about 25% of its maximumtolerated dose following a traditional dosing regime. In someembodiments, the paclitaxel/albumin nanoparticles have an averagediameter of no greater than about 200 nm. In some embodiments, thepaclitaxel/albumin nanoparticle composition is substantially free (suchas free) of surfactant (such as Cremophor). In some embodiments, theweight ratio of the albumin to paclitaxel in the composition is about18:1 or less, such as about 9:1 or less. In some embodiments, thepaclitaxel is coated with albumin. In some embodiments, thepaclitaxel/albumin nanoparticles have an average diameter of no greaterthan about 200 nm and the paclitaxel/albumin composition issubstantially free (such as free) of surfactant (such as Cremophor). Insome embodiments, the paclitaxel/albumin nanoparticles have an averagediameter of no greater than about 200 nm and the paclitaxel is coatedwith albumin. In some embodiments, the nanoparticle composition isAbraxane™.

In some embodiments, there is provided a method of administering acomposition comprising nanoparticles comprising a taxane and a carrierprotein (such as albumin), wherein the nanoparticle composition isadministered over a period of at least one month, wherein the intervalbetween each administration is no more than about a week, and whereinthe dose of the taxane at each administration is about 0.25 mg/m² toabout 25 mg/m². In some embodiments, the taxane is coated with thecarrier protein (such as albumin). In some embodiments, the dose of thetaxane per administration is less than about any of 2, 3, 4, 5, 6, 7, 8,9, 10, 11, 12, 13, 14, 15, 18, 20, 22, and 25 mg/m². In someembodiments, the taxane is administered at least about any of 1×, 2×,3×, 4', 5×, 6×, 7× (i.e., daily) a week. In some embodiments, theintervals between each administration are less than about any of 7 days,6 days, 5 days, 4 days, 3 days, 2 days, and 1 day. In some embodiments,the taxane is administered over a period of at least about any of 2, 3,4, 5, 6, 7, 8, 9, 10, 11, 12, 18, 24, 30 and 36 months.

In some embodiments, there is provided a method of administering acomposition comprising nanoparticles comprising paclitaxel and analbumin, wherein the nanoparticle composition is administered over aperiod of at least one month, wherein the interval between eachadministration is no more than about a week, and wherein the dose of thetaxane at each administration is about 0.25 mg/m² to about 25 mg/m². Insome embodiments, the paclitaxel/albumin nanoparticles have an averagediameter of no greater than about 200 nm. In some embodiments, thepaclitaxel/albumin nanoparticle composition is substantially free (suchas free) of surfactant (such as Cremophor). In some embodiments, theweight ratio of the albumin to paclitaxel in the composition is about18:1 or less, such as about 9:1 or less. In some embodiments, thepaclitaxel is coated with albumin. In some embodiments, thepaclitaxel/albumin nanoparticles have an average diameter of no greaterthan about 200 nm and the paclitaxel/albumin composition issubstantially free (such as free) of surfactant (such as Cremophor). Insome embodiments, the paclitaxel/albumin nanoparticles have an averagediameter of no greater than about 200 nm and the paclitaxel is coatedwith albumin. In some embodiments, the nanoparticle composition isAbraxane™.

In some embodiments, the Abraxane™ (or other paclitaxel/albuminnanoparticle compositions) is administered at the dose of about 3 mg/kgto about 10 mg/kg daily. In some embodiments, the Abraxane™ isadministered at the dose of about 6 mg/kg to about 10 mg/kg daily. Insome embodiments, the Abraxane™ is administered at the dose of about 6mg/kg daily. In some embodiments, Abraxane™ is administered at the doseof about 3 mg/kg daily.

The invention also provides compositions for use in the metronomicregime(s) described herein. In some embodiments, there is provided acomposition comprising nanoparticles comprising a taxane and a carrierprotein (such as albumin), wherein said composition is administered toan individual via a metronomic dosing regime, such as the dosing regimedescribed herein.

OTHER ASPECTS OF THE INVENTION

In another aspects, there are provided methods of treating proliferativediseases comprising administering a composition comprising nanoparticlescomprising a taxane (including pacitiaxel, docetaxel, or ortataxel) anda carrier protein (such as albumin). In some embodiments, there isprovided a method of treating cancer comprising administering acomposition comprising nanoparticles comprising ortataxel and a carrierprotein (such as albumin).

In some embodiments, there is provided methods of treating proliferativediseases comprising administering a composition comprising nanoparticlescomprising a thiocolchicine or its derivative (such as dimericthiocolchicine) and a carrier protein (such as albumin). In someembodiments, there is provided a method of treating cancer comprisingadministering a composition comprising nanoparticles comprising dimericcolchicines and a carrier protein (such as albumin). In someembodiments, the nanoparticle composition is any of (and in someembodiments selected from the group consisting of) Nab-5404, Nab-5800,and Nab-5801.

In some embodiments, there is provided a method of treating cancercomprising administering a composition comprising nanoparticlescomprising paclitaxel, wherein the nanoparticle composition isadministered according to any of the dosing regimes described in Table3. In some embodiments, the cancer is a Taxane refractory metastaticbreast cancer. TABLE 3 Row Study therapy No. Combination Regimen/Dosagetype Protocol title 1. ABX alone ABX: 125 mg/m² qwk × ¾ Metastatic PhaseII study with Breast Cancer weekly Abraxane ™ treatment in taxane-refractory MBC patients 2. ABX alone Arm 1: ABX 130 mg/m² qwk Metastatic3-arm phase II trial in 1st- Arm 2: ABX 260 mg/m² q2wk Breast Cancerline Her-2-MBC patients. Arm 3: ABX 260 mg/m² q3wk 3. ABX alone ABX: 260mg/m² q3wk Metastatic Phase II Controlled, (Capxol) vs Breast CancerRandomized, Open Label Taxol: 175 mg/m² q3wk Study to Evaluate theEfficacy and Safety of Capxol (a Cremophor- Free NanoparticlePaclitaxel) and cremophor-formulated paclitaxel injection in Patientwith Metastatic Breast Cancer 4. ABX alone Arm 1: ABX weekly Metastatic3-arm phase II trial in 1st- Arm 2: ABX q3wk Breast Cancer line and2nd-line MBC, Arm 3: Taxol weekly with biological correlates analysis 5.ABX alone ABX: 300 mg/m² q3wk Stage IIA, IIB, Phase II trial of IIIA,IIIB and neoadjuvant IV breast chemotherapy (NCT) with cancernanoparticle paclitaxel (ABI-007, Abraxane) in women with clinical stageIIA, IIB, IIIA, IIIB and IV (with intact primary) breast cancers 6. ABXalone ABX: 125 mg/m² qwk × ¾ 1st-line Phase I/II study of advancedAbraxane monotherapy in NSCLC 1st-line advanced NSCLC 7. ABX alone ABX260 mg/m² 1st-line NSCLC Phase II ABX mono in q3wk 1st-line NSCLC 8. ABXalone Arm 1: ABX q3wk 2^(nd) line NSCLC Phase II study of Arm 2: ABX qwkAbraxane monotherapy in Doses TBD 2^(nd)-line NSCLC 9. ABX alone ABX:100 mg/m² qwk Prostate Cancer Randomized phase II vs study Abraxane ™weekly ABX: 260 mg/m² q3wk vs every three weeks in front line HRP 10.ABX alone ABX qwk Prostate Cancer Phase II ABX in 1st-line prostatecancer 11. ABX alone ABX: 150 mg/m² qwk × ¾ for 2 Prostate Cancer PhaseII neoadjuvant cycles study 12. ABX alone ABX: 100 mg/m² qwk (no break)Prostate Cancer Phase II ABX 100 mg weekly no break 13. ABX alone ABX:100 mg/m² (previously Malignant Phase II previously treated treated)Melanoma and untreated metastatic ABX: 150 mg/m² (untreated) melanomapatients qwk × ¾ 14. ABX alone ABX: 125 mg/m² Carcinoma of Phase IIstudy of ABX in qwk × ¾ the cervix treatment of persistent or recurrentcarcinoma of the cervix 15. ABX alone Ovarian Cancer Phase II study ofAbraxane for treatment of advanced ovarian cancer (3^(rd) line) 16. ABXalone non-hematologic Phase II single treatment (ABI-007) malignanciesuse of ABI-007 for the treatment of non- hematologic malignancies.Compassionate useNanoparticle Compositions

The nanoparticle compositions described herein comprise nanoparticlescomprising (in various embodiments consisting essentially of) a taxane(such as paclitaxel) and a carrier protein (such as albumin).Nanoparticles of poorly water soluble drugs (such as taxane) have beendisclosed in, for example, U.S. Pat. Nos. 5,916,596; 6,506,405; and6,537,579 and also in U.S. Pat. Pub. No. 2005/0004002A1. Although thedescription provided below is specific to taxane, it is understood thatthe same applies to other drugs, such as rapamycin, 17-AAG, and dimericthiocolchicine.

In some embodiments, the composition comprises nanoparticles with anaverage or mean diameter of no greater than about 1000 nanometers (nm),such as no greater than about any of 900, 800, 700, 600, 500, 400, 300,200, and 100 nm. In some embodiments, the average or mean diameters ofthe nanoparticles is no greater than about 200 nm. In some embodiments,the average or mean diameters of the nanoparticles is no greater thanabout 150 nm. In some embodiments, the average or mean diameters of thenanoparticles is no greater than about 100 nm. In some embodiments, theaverage or mean diameter of the nanoparticles is about 20 to about 400nm. In some embodiments, the average or mean diameter of thenanoparticles is about 40 to about 200 nm. In some embodiments, thenanoparticles are sterile-filterable.

The nanoparticles described herein may be present in a dry formulation(such as lyophilized composition) or suspended in a biocompatiblemedium. Suitable biocompatible media include, but are not limited to,water, buffered aqueous media, saline, buffered saline, optionallybuffered solutions of amino acids, optionally buffered solutions ofproteins, optionally buffered solutions of sugars, optionally bufferedsolutions of vitamins, optionally buffered solutions of syntheticpolymers, lipid-containing emulsions, and the like.

The term “proteins” refers to polypeptides or polymers of amino acids ofany length (including full length or fragments), which may be linear orbranched, comprise modified amino acids, and/or be interrupted bynon-amino acids. The term also encompasses an amino acid polymer thathas been modified naturally or by intervention; for example, disulfidebond formation, glycosylation, lipidation, acetylation, phosphorylation,or any other manipulation or modification. Also included within thisterm are, for example, polypeptides containing one or more analogs of anamino acid (including, for example, unnatural amino acids, etc.), aswell as other modifications known in the art. The proteins describedherein may be naturally occurring, i.e., obtained or derived from anatural source (such as blood), or synthesized (such as chemicallysynthesized or by synthesized by recombinant DNA techniques).

Examples of suitable carrier proteins include proteins normally found inblood or plasma, which include, but are not limited to, albumin,immunoglobulin including IgA, lipoproteins, apolipoprotein B, alpha-acidglycoprotein, beta-2-macroglobulin, thyroglobulin, transferin,fibronectin, factor VII, factor VIII, factor IX, factor X, and the like.In some embodiments, the carrier protein is non-blood protein, such ascasein, α-lactalbumin, and β-lactoglobulin. The carrier proteins mayeither be natural in origin or synthetically prepared. In someembodiments, the pharmaceutically acceptable carrier comprises albumin,such as human serum albumin. Human serum albumin (HSA) is a highlysoluble globular protein of M_(r) 65K and consists of 585 amino acids.HSA is the most abundant protein in the plasma and accounts for 70-80%of the colloid osmotic pressure of human plasma. The amino acid sequenceof HSA contains a total of 17 disulphide bridges, one free thiol (Cys34), and a single tryptophan (Trp 214). Intravenous use of HSA solutionhas been indicated for the prevention and treatment of hypovolumic shock(see, e.g.; Tullis, JAMA, 237, 355-360, 460-463, (1977)) and Houser etal., Surgery, Gynecology and Obstetrics, 150, 811-816 (1980)) and inconjunction with exchange transfusion in-the treatment of neonatalhyperbilirubinemia (see, e.g., Finlayson, Seminars in Thrombosis andHemostasis, 6, 85-120, (1980)). Other albumins are contemplated, such asbovine serum albumin. Use of such non-human albumins could beappropriate, for example, in the context of use of these compositions innon-human mammals, such as the veterinary (including domestic pets andagricultural context).

Human serum albumin (HSA) has multiple hydrophobic binding sites (atotal of eight for fatty acids, an endogenous ligand of HSA) and binds adiverse set of taxanes, especially neutral and negatively chargedhydrophobic compounds (Goodman et al., The Pharmacological Basis ofTherapeutics, 9^(th) ed, McGraw-Hill New York (1996)). Two high affinitybinding sites have been proposed in subdomains IIA and IIIA of HSA,which are highly elongated hydrophobic pockets with charged lysine andarginine residues near the surface which function as attachment pointsfor polar ligand features (see, e.g., Fehske et al., Biochem. Pharmcol,30, 687-92 (198a), Vorum, Dan. Med. Bull., 46, 379-99 (1999),Kragh-Hansen, Dan. Med. Bull., 1441, 131-40 (1990), Curry et al., Nat.Struct. Biol., 5, 827-35 (1998), Sugio et al., Protein. Eng., 12, 439-46(1999), He et al., Nature, 358, 209-15 (199b), and Carter et al., Adv.Protein. Chem., 45, 153-203 (1994)). Paclitaxel and propofol have beenshown to bind HSA (see, e.g., Paal et al., Eur. J. Biochem., 268(7),2187-91 (200a), Purcell et al., Biochim. Biophys. Acta, 1478(a), 61-8(2000), Altmayer et al., Arzneimittelforschung, 45, 1053-6 (1995), andGarrido et al., Rev. Esp. Anestestiol. Reanim., 41, 308-12 (1994)). Inaddition, docetaxel has been shown to bind to human plasma proteins(see, e.g., Urien et al., Invest. New Drugs, 14(b), 147-51 (1996)).

The carrier protein (such as albumin) in the composition generallyserves as a carrier for the taxane, i.e., the carrier protein in thecomposition makes the taxane more readily suspendable in an aqueousmedium or helps maintain the suspension as compared to compositions notcomprising a carrier protein. This can avoid the use of toxic solvents(or surfactants) for solubilizing the taxane, and thereby can reduce oneor more side effects of administration of the taxane into an individual(such as a human). Thus, in some embodiments, the composition describedherein is substantially free (such as free) of surfactants, such asCremophor (including Cremophor EL® (BASF)). In some embodiments, thenanoparticle composition is substantially free (such as free) ofsurfactants. A composition is “substantially free of Cremophor” or“substantially free of surfactant” if the amount of Cremophor orsurfactant in the composition is not sufficient to cause one or moreside effect(s) in an individual when the nanoparticle composition isadministered to the individual.

The amount of carrier protein in the composition described herein willvary depending on other components in the composition. In someembodiments, the composition comprises a carrier protein in an amountthat is sufficient to stabilize the taxane in an aqueous suspension, forexample, in the form of a stable colloidal suspension (such as a stablesuspension of nanoparticles). In some embodiments, the carrier proteinis in an amount that reduces the sedimentation rate of the taxane in anaqueous medium. For particle-containing compositions, the amount of thecarrier protein also depends on the size and density of nanoparticles ofthe taxane.

A taxane is “stabilized” in an aqueous suspension if it remainssuspended in an aqueous medium (such as without visible precipitation orsedimentation) for an extended period of time, such as for at leastabout any of 0.1, 0.2, 0.25, 0.5, 1, 2, 3, 4, 5, 6, 7, 8, 9, 10, 11, 12,24, 36, 48, 60, or 72 hours. The suspension is generally, but notnecessarily, suitable for administration to an individual (such ashuman). Stability of the suspension is generally (but not necessarily)evaluated at a storage temperature (such as room temperature (such as20-25° C.) or refrigerated conditions (such as 4° C.)). For example, asuspension is stable at a storage temperature if it exhibits noflocculation or particle agglomeration visible to the naked eye or whenviewed under the optical microscope at 1000 times, at about fifteenminutes after preparation of the suspension. Stability can also beevaluated under accelerated testing conditions, such as at a temperaturethat is higher than about 40° C.

In some embodiments, the carrier protein is present in an amount that issufficient to stabilize the taxane in an aqueous suspension at a certainconcentration. For example, the concentration of the taxane in thecomposition is about 0.1 to about 100 mg/ml, including for example anyof about 0.1 to about 50 mg/ml, about 0.1 to about 20 mg/ml, about 1 toabout 10 mg/ml, about 2 mg/ml to about 8 mg/ml, about 4 to about 6mg/ml, about 5 mg /ml. In some embodiments, the concentration of thetaxane is at least about any of 1.3 mg/ml, 1.5 mg/ml, 2 mg/ml, 3 mg/ml,4 mg/ml, 5 mg/ml, 6 mg/ml, 7 mg/ml, 8 mg/ml, 9 mg/ml, 10 mg/ml, 15mg/ml, 20 mg/ml, 25 mg/ml, 30 mg/ml, 40 mg/ml, and 50 mg/ml. In someembodiments, the carrier protein is present in an amount that avoids useof surfactants (such as Cremophor), so that the composition is free orsubstantially free of surfactant (such as Cremophor).

In some embodiments, the composition, in liquid form, comprises fromabout 0.1% to about 50% (w/v) (e.g. about 0.5% (w/v), about 5% (w/v),about 10% (w/v), about 15% (w/v), about 20% (w/v), about 30% (w/v),about 40% (w/v), or about 50% (w/v)) of carrier protein. In someembodiments, the composition, in liquid form, comprises about 0.5% toabout 5% (w/v) of carrier protein.

In some embodiments, the weight ratio of carrier protein, e.g., albumin,to the taxane in the nanoparticle composition is such that a sufficientamount of taxane binds to, or is transported by, the cell. While theweight ratio of carrier protein to taxane will have to be optimized fordifferent carrier protein and taxane combinations, generally the weightratio of carrier protein, e.g., albumin, to taxane (w/w) is about 0.01:1to about 100: 1, about 0.02:1 to about 50:1, about 0.05:1 to about 20:1,about 0.1:1 to about 20:1, about 1:1 to about 18:1, about 2:1 to about15:1, about 3:1 to about 12:1, about 4:1 to about 10:1, about 5:1 toabout 9:1, or about 9:1. In some embodiments, the carrier protein totaxane weight ratio is about any of 18:1 or less, 15:1 or less, 14:1 orless, 13:1 or less, 12:1 or less, 11:1 or less, 10:1 or less, 9:1 orless, 8:1 or less, 7:1 or less, 6:1 or less, 5:1 or less, 4:1 or less,and 3:1 or less.

In some embodiments, the carrier protein allows the composition to beadministered to an individual (such as human) without significant sideeffects. In some embodiments, the carrier protein (such as albumin) isin an amount that is effective to reduce one or more side effects ofadministration of the taxane to a human. The term “reducing one or moreside effects of administration of the taxane” refers to reduction,alleviation, elimination, or avoidance of one or more undesirableeffects caused by the taxane, as well as side effects caused by deliveryvehicles (such as solvents that render the taxanes suitable forinjection) used to deliver the taxane. Such side effects include, forexample, myelosuppression, neurotoxicity, hypersensitivity,inflammation, venous irritation, phlebitis, pain, skin irritation,peripheral neuropathy, neutropenic fever, anaphylactic reaction, venousthrombosis, extravasation, and combinations thereof. These side effects,however, are merely exemplary and other side effects, or combination ofside effects, associated with taxanes can be reduced.

In some embodiments, the composition comprises Abraxane™. Abraxane™ is aformulation of paclitaxel stabilized by human albumin USP, which can bedispersed in directly injectable physiological solution. When dispersedin a suitable aqueous medium such as 0.9% sodium chloride injection or5% dextrose injection, Abraxane™ forms a stable colloidal suspension ofpaclitaxel. The mean particle size of the nanoparticles in the colloidalsuspension is about 130 nanometers. Since HSA is freely soluble inwater, Abraxane™ can be reconstituted in a wide range of concentrationsranging from dilute (0.1 mg/ml paclitaxel) to concentrated (20 mg/mlpaclitaxel), including for example about 2 mg/ml to about 8 mg/ml, about5 mg/ml.

Methods of making nanoparticle compositions are known in the art. Forexample, nanoparticles containing taxanes (such as paclitaxel) andcarrier protein (such as albumin) can be prepared under conditions ofhigh shear forces (e.g., sonication, high pressure homogenization, orthe like). These methods are disclosed in, for example, U.S. Pat. Nos.5,916,596; 6,506,405; and 6,537,579 and also in U.S. Pat. Pub. No.2005/0004002A1.

Briefly, the taxane (such as docetaxel) is dissolved in an organicsolvent, and the solution can be added to a human serum albuminsolution. The mixture is subjected to high pressure homogenization. Theorganic solvent can then be removed by evaporation. The dispersionobtained can be further lyophilized. Suitable organic solvent include,for example, ketones, esters, ethers, chlorinated solvents, and othersolvents known in the art. For example, the organic solvent can bemethylene chloride and chloroform/ethanol (for example with a ratio of1:9, 1:8, 1:7, 1:6, 1:5, 1:4, 1:3, 1:2, 1:1, 2:1, 3:1, 4:1, 5:1, 6:1,7:1, 8:1, or 9:a).

Other Components in the Nanoparticle Compositions

The nanoparticles described herein can be present in a composition thatinclude other agents, excipients, or stabilizers. For example, toincrease stability by increasing the negative zeta potential ofnanoparticles, certain negatively charged components may be added. Suchnegatively charged components include, but are not limited to bile saltsof bile acids consisting of glycocholic acid, cholic acid,chenodeoxycholic acid, taurocholic acid, glycochenodeoxycholic acid,taurochenodeoxycholic acid, litocholic acid, ursodeoxycholic acid,dehydrocholic acid and others; phospholipids including lecithin (eggyolk) based phospholipids which include the followingphosphatidylcholines: palmitoyloleoylphosphatidylcholine,palmitoyllinoleoylphosphatidylcholine,stearoyllinoleoylphosphatidylcholine stearoyloleoylphosphatidylcholine,stearoylarachidoylphosphatidylcholine, anddipalmitoylphosphatidylcholine. Other phospholipids includingL-α-dimyristoylphosphatidylcholine (DMPC), dioleoylphosphatidylcholine(DOPC), distearyolphosphatidylcholine (DSPC), hydrogenated soyphosphatidylcholine (HSPC), and other related compounds. Negativelycharged surfactants or emulsifiers are also suitable as additives, e.g.,sodium cholesteryl sulfate and the like.

In some embodiments, the composition is suitable for administration to ahuman. In some embodiments, the composition is suitable foradministration to a mammal such as, in the veterinary context, domesticpets and agricultural animals. There are a wide variety of suitableformulations of the nanoparticle composition (see, e.g., U.S. Pat. Nos.5,916,596 and 6,096,331). The following formulations and methods aremerely exemplary and are in no way limiting. Formulations suitable fororal administration can consist of (a) liquid solutions, such as aneffective amount of the compound dissolved in diluents, such as water,saline, or orange juice, (b) capsules, sachets or tablets, eachcontaining a predetermined amount of the active ingredient, as solids orgranules, (c) suspensions in an appropriate liquid, and (d) suitableemulsions. Tablet forms can include one or more of lactose, mannitol,corn starch, potato starch, microcrystalline cellulose, acacia, gelatin,colloidal silicon dioxide, croscarmellose sodium, talc, magnesiumstearate, stearic acid, and other excipients, colorants, diluents,buffering agents, moistening agents, preservatives, flavoring agents,and pharmacologically compatible excipients. Lozenge forms can comprisethe active ingredient in a flavor, usually sucrose and acacia ortragacanth, as well as pastilles comprising the active ingredient in aninert base, such as gelatin and glycerin, or sucrose and acacia,emulsions, gels, and the like containing, in addition to the activeingredient, such excipients as are known in the art.

Examples of suitable carriers, excipients, and diluents include, but arenot limited to, lactose, dextrose, sucrose, sorbitol, mannitol,starches, gum acacia, calcium phosphate, alginates, tragacanth, gelatin,calcium silicate, microcrystalline cellulose, polyvinylpyrrolidone,cellulose, water, saline solution, syrup, methylcellulose, methyl- andpropylhydroxybenzoates, talc, magnesium stearate, and mineral oil. Theformulations can additionally include lubricating agents, wettingagents, emulsifying and suspending agents, preserving agents, sweeteningagents or flavoring agents.

Formulations suitable for parenteral administration include aqueous andnon-aqueous, isotonic sterile injection solutions, which can containanti-oxidants, buffers, bacteriostats, and solutes that render theformulation compatible with the blood of the intended recipient, andaqueous and non-aqueous sterile suspensions that can include suspendingagents, solubilizers, thickening agents, stabilizers, and preservatives.The formulations can be presented in unit-dose or multi-dose sealedcontainers, such as ampules and vials, and can be stored in afreeze-dried (lyophilized) condition requiring only the addition of thesterile liquid excipient, for example, water, for injections,immediately prior to use. Extemporaneous injection solutions andsuspensions can be prepared from sterile powders, granules, and tabletsof the kind previously described. Injectable formulations are referred.

In some embodiments, the composition is formulated to have a pH range ofabout 4.5 to about 9.0, including for example pH ranges of any of about5.0 to about 8.0, about 6.5 to about 7.5, and about 6.5 to about 7.0. Insome embodiments, the pH of the composition is formulated to no lessthan about 6, including for example no less than about any of 6.5, 7, or8 (such as about 8). The composition can also be made to be isotonicwith blood by the addition of a suitable tonicity modifier, such asglycerol.

Kits

The invention also provides kits for use in the instant methods. Kits ofthe invention include one or more containers comprisingtaxane-containing nanoparticle compositions (or unit dosage forms and/orarticles of manufacture) and/or a chemotherapeutic agent, and in someembodiments, further comprise instructions for use in accordance withany of the methods described herein. The kit may further comprise adescription of selection an individual suitable or treatment.Instructions supplied in the kits of the invention are typically writteninstructions on a label or package insert (e.g., a paper sheet includedin the kit), but machine-readable instructions (e.g., instructionscarried on a magnetic or optical storage disk) are also acceptable.

In some embodiments, the kit comprises a) a composition comprisingnanoparticles comprising a taxane and a carrier protein (such asalbumin), b) an effective amount of at least one other chemotherapeuticagent, and c) instructions for administering the nanoparticles and thechemotherapeutic agents simultaneously and/or sequentially, fortreatment of a proliferative disease (such as cancer). In someembodiments, the taxane is any of paclitaxel, docetaxel, and ortataxel.In some embodiments, the kit comprises nanoparticles comprising a) acomposition comprising nanoparticles comprising paclitaxel and analbumin (such as Abraxane™), b) an effective amount of at least oneother chemotherapeutic agent, and c) instructions for administering thenanoparticles and the chemotherapeutic agents simultaneously and/orsequentially, for the effective treatment of a proliferative disease(such as cancer).

In some embodiments, the kit comprises a) a composition comprisingnanoparticles comprising a taxane and a carrier protein (such asalbumin), b) a composition comprising nanoparticles comprising at leastone other chemotherapeutic agent and a carrier protein (such asalbumin), and c) instructions for administering the nanoparticlecompositions simultaneously and/or sequentially, for treatment of aproliferative disease (such as cancer). In some embodiments, the kitcomprises nanoparticles comprising a) a composition comprisingnanoparticles comprising paclitaxel and an albumin (such as Abraxane™),b) a composition comprising nanoparticles comprising at least one otherchemotherapeutic agent and a carrier protein (such as albumin), and c)instructions for administering the nanoparticle compositionssimultaneously and/or sequentially, for the effective treatment of aproliferative disease (such as cancer).

The nanoparticles and the chemotherapeutic agents can be present inseparate containers or in a single container. It is understood that thekit may comprise one distinct composition or two or more compositionswherein one composition comprises nanoparticles and one compositioncomprises a chemotherapeutic agent.

The kits of the invention are in suitable packaging. Suitable packaginginclude, but is not limited to, vials, bottles, jars, flexible packaging(e.g., seled Mylar or plastic bags), and the like. Kits may optionallyprovide additional components such as buffers and interpretativeinformation.

The instructions relating to the use of the nanoparticle compositionsgenerally include information as to dosage, dosing schedule, and routeof administration for the intended treatment. The containers may be unitdoses, bulk packages (e.g., multi-dose packages) or sub-unit doses. Forexample, kits may be provided that contain sufficient dosages of thetaxane (such as taxane) as disclosed herein to provide effectivetreatment of an individual for an extended period, such as any of aweek, 2 weeks, 3 weeks, 4 weeks, 6 weeks, 8 weeks, 3 months, 4 months, 5months, 7 months, 8 months, 9 months, or more. Kits may also includemultiple unit doses of the taxane and pharmaceutical compositions andinstructions for use and packaged in quantities sufficient for storageand use in pharmacies, for example, hospital pharmacies and compoundingpharmacies.

Those skilled in the art will recognize that several variations arepossible within the scope and spirit of this invention. The inventionwill now be described in greater detail by reference to the followingnon-limiting examples. The following examples further illustrate theinvention but, of course, should not be construed as in any way limitingits scope.

EXAMPLES Example 1 Improved Response and Reduced Toxicities forAbraxane™ Compared to Taxol® in a Phase III Study of Abraxane™ GivenEvery Three Weeks

Significantly reduced incidence of neutropenia and hypersensitivity,absence of requirement of steroid premedication, shorter duration ofneuropathy, shorter infusion time and higher-dose.

ABI-007 (Abraxane™), the first biologically interactive albumin-boundpaclitaxel in a nanoparticle form, free of any solvent, was comparedwith Cremophor®-based paclitaxel (Taxol®) in individuals with metastaticbreast cancer (MBC). This phase III study was performed to confirm thepreclinical studies demonstrating superior efficacy and reduced toxicityof ABI-007 when compared with Taxol®. Individuals were randomly assignedto 3-week cycles of either ABI-007 260 mg/m² (iv) over 30 minuteswithout premedication (n=229) or Taxol® 175 mg/m² IV over 3 hours withpremedication (n=225). ABI-007 demonstrated significantly higherresponse rates compared with Taxol® (33% vs. 19%; p=0.001) andsignificantly longer time to tumor progression (23.0 vs. 16.9 weeks;HR=0.75; p=0.006). There was a trend for longer overall survival inindividuals who received ABI-007 (65.0 vs. 55.7 weeks; p=0.374). In anunplanned analysis, ABI-007 improved survival in individuals receivingtreatment as second- or greater-line therapy (56.4 vs. 46.7 weeks;HR=0.73; p=0.024). The incidence of grade 4 neutropenia wassignificantly lower in the ABI-007 group (9% vs. 22%; p<0.001) despite a49% higher paclitaxel dose. Grade 3 sensory neuropathy was more commonin the ABI-007 group than in the Taxol® group (10% vs. 2%; p<0.001) butwas easily managed and improved more rapidly (median, 22 days) than forTaxol® (median 73 days). No severe (grade 3 or 4) treatment-relatedhypersensitivity reactions occurred in any of the individuals in theABI-007 group despite the absence of premedication and shorteradministration time. In contrast, grade 3 hypersensitivity reactionsoccurred in the Taxol® group despite standard premedication (chest pain:2 individuals; allergic reaction: 3 individuals). Per protocol,corticosteroids and antihistamines were not administered routinely toindividuals in the ABI-007 group; however, premedication wasadministered for emesis, myalgia/arthralgia, or anorexia in 18individuals (8%) in the ABI-007 group in 2% of the treatment cycles,whereas 224 individuals (>99%) in the Taxol® group receivedpremedication at 95% of the cycles. The only clinical chemistry valuethat was notably different between the 2 treatment arms was higher serumglucose levels in the Taxol®-treated individuals, who also had a higherincidence of hyperglycemia reported as an AE (adverse effects) (15 [7%]vs. 3 [1%]; p=0.003). Overall, ABI-007 demonstrated greater efficacy anda favorable safety profile compared with Taxol® in this individualpopulation. The improved therapeutic index and elimination of thesteroid premedication required for solvent-based taxanes make thisnanoparticle albumin-bound paclitaxel an important advance in thetreatment of MBC.

Example 2 Weekly Abraxane™ in Taxane-Refractory Metastatic Breast CancerIndividuals

A recent Phase II clinical study showed that weekly administration ofAbraxane™ (nanoparticle albumin-bound paclitaxel) at a dose of 125 mg/m²resulted in long-term disease control in individuals with metastaticbreast cancer whose disease had progressed while being treated withTaxol® or Taxotere® (that is, individuals who are taxane-refractory).

Abraxane™ is believed to represent the first biologically interactivecomposition that exploits the receptor-mediated (gp60) pathway found tobe integral to achieving high intracellular tumor concentrations of theactive ingredient—paclitaxel. The Phase II study included 75 individualswith taxane-refractory metastatic breast cancer. Abraxane™ wasadministered weekly via a 30-minute infusion at 125 mg/m² withoutsteroid/antihistamine premedication or G-CSF prophylaxis. Individualsreceived three weekly doses followed by one week of rest, repeated every28 days. Unlike Taxol® or Taxotere®, which contain detergents that mayinhibit tumor uptake, the mechanism of action of the albumin-boundnanoparticle paclitaxel may result in improved outcomes, especially inthis difficult-to-treat individual population.

Specifically, the data showed that despite this high weekly dose of 125mg/m² in this highly pre-treated and prior taxane-exposed individualpopulation, only 3 of 75 individuals (4%) had to discontinue Abraxane™due to peripheral neuropathy. Furthermore, of those who experiencedGrade 3 peripheral neuropathy, 80% were typically able to resumetreatment after a delay of only 1 or 2 weeks and continued to receiveAbraxane™ at a reduced dose for an average of 4 additional months. Thisrapid improvement was consistent with our observation from the Phase IIItrial—that the peripheral neuropathy induced by paclitaxel alone (i.e.,without Cremophor®) improves rapidly as compared to that induced byTaxol®. These Abraxane™ clinical trial experiences provide the firstclinical opportunity to evaluate the effects of the chemotherapeuticagent itself, paclitaxel, from the effects from those of solvents. Basedupon both the Phase II and III experience, the data now suggest that theperipheral neuropathy from Abraxane™ is not comparable to the peripheralneuropathy from Taxol® or Taxotere® with respect to duration and impacton the individual.

With regard to the clinical experience of peripheral neuropathyfollowing Taxol® or Taxotere®, Abraxis Oncology recently completed asurvey of 200 oncologists who were asked how long they thought theperipheral neuropathy induced by Taxol® took to improve and/or resolve:25% reported “7-12 months” and another 23% reported “never resolved”;for Taxotere®, the respective percentages were 29% and 7%. These dataare consistent with the statements in the Taxotere® and Taxol® packageinserts.

Analysis of the Phase II data demonstrates Abraxane™ to be active inthis poor-prognosis individual population (87% visceral (lung and liver)disease, 69%>3 metastatic sites, 88% tumor growth while on taxanes), oftaxane-refractory individuals with metastatic breast cancer.Observations included a 44% disease control in Taxotere®-refactoryindividuals and 39% disease control in Taxolo-refractory individuals. Ofthose individuals whose disease progressed while on Taxotereo alone inthe metastatic setting (n=27) a 19% response rate was noted afterreceiving weekly Abraxane™, Of those individuals whose diseaseprogressed while on Taxol® alone in the metastatic setting (n=23) a 13%response rate was noted after receiving weekly Abraxane™.

Abraxane™ was found to be well tolerated when administered weekly over30 minutes without steroids or G-CSF prophylaxis: Grade 4 neutropenia=3%(without G-CSF); Grade 4 anemia=1%; no severe hypersensitivity reactions(despite absence of premedication). In this heavily pretreatedindividual population, 75% of individuals were treated at the full highdose of 125 mg/m² weekly Abraxane™, with no dose reductions due totoxicities/adverse events. Of the individuals who developed grade 3sensory neuropathy, 77% were able to restart Abraxane™ at a reduced dose(75-100 mg/m²) and received a mean of 12.2 (range, 1-28) additionaldoses of Abraxane™. It was remarkable to note that of these individualswho resumed Abraxane™, 80% (8 of 10) were able to restart the drugwithin 14 days after improvement of neuropathy to Grade 1 or 2. Theseresults support the observations in the pivotal Phase III trial of 260mg/m² Abraxane™ administered every 3 weeks, in which rapid improvementof neuropathy (median of 22 days) was also noted. Taken together thesetwo clinical trials suggest when paclitaxel is given alone, theneuropathy which occurs appears to be short-lived and is easily managed.

Abraxane™ utilizes the gp60 receptor based pathway on the microvesselendothelial cells to transport the albumin-paclitaxel complex out of theblood vessel and into the tumor interstitium, and it has been shown thatTaxol® was not transported by this mechanism. Furthermore, analbumin-binding protein, SPARC, was over-expressed in breast tumors andmay play a role in the increased intra-tumoral accumulation ofAbraxane™. The proposed mechanism suggested that once in the tumorinterstitium, the albumin-paclitaxel complex would bind to SPARC thatwas present on the tumor cell surface and be rapidly internalized intothe tumor cell by a non-lysosomal mechanism.

In addition, the surfactants/solvents commonly used in current taxaneformulations such as Cremophor®, Tween® 80 and TPGS, strongly inhibitthe binding of paclitaxel to albumin, thereby limiting transendothelialtransport. Additional data presented showed a statistically improvedefficacy of Abraxane™ over Taxotere® in the MX-1 mammary breastcarcinoma xenograft at equal dose.

In conclusion, 75% of individuals were treated at full high dose with nodose reductions. Data indicate rapid improvement of peripheralneuropathy when nanoparticle albumin-bound paclitaxel is administeredalone, without the solvent Cremophor®. Additional data provide increasedevidence that mechanism of action may play important role in enhancingindividual outcomes.

Example 3 Abraxane™ (ABI-007) Acts Synergistically with TargetedAntiangiogenic Pro-Apoptotic Peptides (HKP) in MDA-MB-435 Human TumorXenografts

The antiangiogenic activity of small synthetic pro-apoptotic peptidescomposed of two functional domains, one targeting the CD13 receptors(aminopeptidase N) on tumor microvessels and the other disrupting themitochondrial membrane following internalization have previously beenreported. See Nat Med. September; 1999 5(9):1.032-8. A second generationdimeric peptide, CNGRC-GG-d(KLAKLAK)₂, named HKP (Hunter Killer Peptide)was found to have improved antitumor activity. Since anti-angiogenicagents such as Avastin® exhibit synergism in combination with cytotoxicagents such as 5-fluorouracil, we evaluated the combination of theantiangiogenic HKP with Abraxane™ (ABI-007), an albumin nanoparticlepaclitaxel that is transported by the gp60 receptor in vascularendothelium (Desai, SABCS-2003), in MDA-MB-435 human breast tumorxenografts.

Methods: MDA-MB-435 human tumor xenografts were established at anaverage tumor volume of 100 mm³, mice were randomized into groups of12-13 animals and treated with HKP, Abraxane™, or HKP and Abraxane™, HKPwas delivered i.v. (250 ug), once a week, for 16 weeks. Abraxane™ wasadministered i.v., daily for 5 days at 10 mg/kg/day only for the firstweek of treatment. The Abraxane™ dose used was substantially below itsMTD (30 mg/kg/day, qd×5) to prevent the tumor from complete regressionso effect of HKP could be noted.

Results: At nineteen weeks of treatment, tumor volume was significantlydecreased between control group (10,298 mm³±2,570) and HKP (4,372mm³±2,470; p<0.05 vs control) or ABI-007 (3,909 mm³±506; p<0.01 vscontrol). The combination of ABI-007 and HKP significantly reduced thetumor volume over either monotherapy (411 mm³±386; p<0.01 vs. Abraxane™monotherapy or HKP monotherapy). The treatments were well tolerated.

Conclusion: The combination of Abraxane™ (ABI-007), a nanoparticlealbumin-bound paclitaxel, with the vascular targeting anti-angiogenicdimeric peptide HKP (CNGRC-GG-d(KLAKLAK)₂) against the MDA-MB-435xenograft breast tumor showed a significant reduction in tumor volumecompared to monotherapy of either agent alone. Our results suggest thatthe combination of Abraxane™ with antiangiogenic agents such as HKPs orperhaps Avastin® may be beneficial.

Example 4 Metronomic ABI-007 Therapy: Antiangiogenic and AntitumorActivity of a Nanoparticle Albumin-bound Paclitaxel Example 4a

Methods: The antiangiogenic activity of ABI-007 was assessed by the rataortic ring, human umbilical vein endothelial cell (HUVEC) proliferationand tube formation assays. Optimal dose of ABI-007 for metronomictherapy was determined by measuring the levels of circulatingendothelial progenitors (CEPs) in peripheral blood of Balb/c non-tumorbearing mice (n=5/group; dosing: 1-30 mg/kg, i.p, qd×7) with flowcytometry (Shaked et al., Cancer Cell, 7:101-111 (2005)). Subsequently,the antitumor effects of metronomic (qd; i.p.) and MTD (qd×5, 1 cycle;i.v.) ABI-007 and Taxol® were evaluated and compared in SCID micebearing human MDA-MD-231 breast and PC3 prostate cancer xenografts.

Results: ABI-007 at 5 nM significantly (p<0.05) inhibited rat aorticmicrovessel outgrowth, human endothelial cell proliferation and tubeformation by 53%, 24%, and 75%, respectively. The optimal dose ofABI-007 for metronomic therapy was observed to be 6-10. mg/kg based onCEP measurements. Metronomic ABI-007 (6 mg/kg) but not Taxol® (1.3mg/kg) significantly (p<0.05) suppressed tumor growth in both xenograftmodels. Neither ABI-007 nor Taxol® administered metronomically inducedany weight loss. Although MTD ABI-007 (30 mg/kg) inhibited tumor growthmore effectively than MTD Taxol® (13 mg/kg), significant weight loss wasnoted with the former. Interestingly, the antitumor effect of metronomicABI-007 approximated that of MTD Taxol®.

Conclusion: ABI-007 exhibits potent antiangiogenic and antitumoractivity when used in a metronomic regime.

Example 4b

Rat Aortic Ring Assay. Twelve-well tissue culture plates were coatedwith Matrigel (Collaborative Biomedical Products, Bedford, Mass.) andallowed to gel for 30 min at 37° C. and 5% CO₂. Thoracic aortas wereexcised from 8- to 10-week-old male Sprague-Dawley rats, cut into1-mm-long cross-sections, placed on Matrigel-coated wells and coveredwith an additional Matrigel. After the second layer of Matrigel had set,the rings were covered with EGM-II and incubated overnight at 37° C. and5% CO₂. EGM-II consists of endothelial cell basal medium (EBM-II;Cambrex, Walkersville, Md.) plus endothelial cell growth factorsprovided as the EGM-II Bulletkit (Cambrex). The culture medium wassubsequently changed to EBM-II supplemented with 2% FBS, 0.25 μg/mlamphotericin B and 10 μg/ml gentamycin. Aortic rings were treated withEBM-II containing the vehicle (0.9% saline/albumin),carboxyamidotriazole (CAI; 12 μg/ml), or ABI-007 (0.05-10 nM paclitaxel)for 4 days and photographed on the fifth day. CAI, a knownanti-angiogenic agent, was used at a higher than clinically achievableconcentration as a positive control. Experiments were repeated fourtimes using aortas from four different rats. The area of angiogenicsprouting, reported in square pixels, was quantified using AdobePhotoshop 6.0.

As shown in FIG. 1A, ABI-007 significantly inhibited rat aorticmicrovessel outgrowth in a concentration-dependent manner relative tothe vehicle control, reaching statistical significance (p<0.05) at 5 nM(53% inhibition) and 10 nM (68% inhibition). The amount of albuminpresent at each concentration of ABI-007 alone did not inhibitangiogenesis.

Endothelial Cell Proliferation Assay. Human umbilical vein endothelialcells (HUVEC; Cambrex) were maintained in EGM-II at 37° C. and 5% CO2.HUVECs were seeded onto 12-well plates at a density of 30,000 cells/welland allowed to attach overnight. The culture medium was then aspirated,and fresh culture medium containing either the vehicle (0.9%saline/albumin), or ABI-007 (0.05-10 nM paclitaxel) was added to eachwell. After 48 h, cells were trypsinized and counted with a Coulter Z1counter (Coulter Corp., Hialeah, Fla.). All experiments were repeatedthree times.

As shown in FIG. 1B, human endothelial cell proliferation wassignificantly inhibited by ABI-007 at 5 nM and 10 nM by 36% and 41%,respectively.

Endothelial Cell Tube Formation Assay. Eight-well slide chambers werecoated with Matrigel and allowed to gel at 37° C. and 5% CO₂ for 30 min.HUVECs were then seeded at 30,000 cells/well in EGM-II containing eitherthe vehicle (0.9% saline/albumin) or ABI-007 (0.05-10 nM paclitaxel) andincubated at 37° C. and 5% CO₂ for 16 h. After incubation, slides werewashed in PBS, fixed in 100% methanol for 10 s, and stained withDiffQuick solution II (Dade Behring Inc., Newark, Del.) for 2 min. Toanalyze tube formation, each well was digitally photographed using a2.5× objective. A threshold level was set to mask the stained tubes. Thecorresponding area was measured as the number of pixels using MetaMorphsoftware (Universal Imaging, Downingtown, Pa.). Experiments wererepeated three times.

As shown in FIG. 1C, ABI-007 blocked tube formation by 75% at both 5 nMand 10 nM.

Determination of the In Vivo Optimal Biologic Dose of ABI-007 byMeasuring Circulating Endothelial Cells (CECs) and CirculatingEndothelial Progenitors (CEPs). Six- to eight-week-old female Balb/cJmice were randomized into the following eight groups (n=5 each):untreated, treated with i.p. bolus injections of either the drug vehicle(0.9% saline/albumin), or ABI-007 at 1, 3, 6, 10, 15 or 30 mg/kgpaclitaxel daily for 7 days. At the end of the treatment period, bloodsamples were drawn by cardiac puncture and collected in EDTA-containingvacutainer tubes (Becton Dickinson, Franklin Lakes, N.J.). CECs and CEPswere enumerated using four-color flow cytometry. Monoclonal antibodiesspecific for CD45 were used to exclude CD45+ hematopoietic cells. CECsand their CEP subset were depicted using the murine endothelial markersfetal liver kinase 1/VEGF receptor 2 (flk-1/VEGFR2), CD13, and CD117 (BDPharmingen, San Diego, Calif.). Nuclear staining (Procount; BDBiosciences, San Jose, Calif.) was performed to exclude the possibilityof platelets or cellular debris interfering with the accuracy of CEC andCEP enumeration. After red cell lysis, cell suspensions were evaluatedby a FACSCalibur (BD Biosciences) using analysis gates designed toexclude dead cells, platelets, and debris. At least 100,000events/sample were obtained in order to analyze the percentage of CECsand CEPs. The absolute number of CECs and CEPs was then calculated asthe percentage of the events collected in the CEC and CEP enumerationgates multiplied by the total white cell count. Percentages of stainedcells were determined and compared to the appropriate negative controls.Positive staining was defined as being greater than non-specificbackground staining. 7-aminoactinomycin D (7AAD) was used to enumerateviable versus apoptotic and dead cells.

FIG. 2 shows that ABI-007 administered i.p. daily for 7 days at 3, 10-30mg/kg significantly decreased CEP levels in non-tumor bearing Balb/cJmice. However, ABI-007 at 10-30 mg/kg was associated with a significantreduction of white blood cell count indicative of toxicity. Although thereduction of CEP levels by ABI-007 at 6 mg/kg did not reach statisticalsignificance, decrease in white blood cell count was not evident.Therefore it was concluded that the in vivo optimal biologic dose formetronomic ABI-007 was between 3-10 mg/kg. In one study, metronomicTaxol® at 1.3, 3, 6, or 13 mg/kg given i.p. daily for 7 days did notsignificantly reduce viable CEP levels, whereas metronomic Taxol® at 30mg/kg or higher resulted in severe toxicity and eventually mortality inmice. It was previously reported that the i.p. administration of Taxol®at doses commonly used in the clinic resulted in entrapment ofpaclitaxel in Cremophor® EL micelles in the peritoneal cavity andconsequently, insignificant plasma paclitaxel concentration (Gelderblomet al., Clin. Cancer Res. 8:1237-41 (2002)). This would explain whydoses of metronomic Taxol® (1.3, 3, 6, and 13 mg/kg) that did not causedeath failed to change viable CEP levels. In this case, the i.p.administration of metronomic Taxol® at 1.3 mg/kg would not be anydifferent from that at 13 mg/kg. Therefore the lower dose, 1.3 mg/kg,was selected to minimize the amount of Cremophor® EL per paclitaxeladministration for subsequent experiments.

Antitumor effects of metronomic and MTD ABI-007 compared with metronomicand MTD Taxol®. Human prostate cancer cell line PC3 and human breastcancer cell line MDA-MD-231 were obtained from the American Type CultureCollection (Manassas, Va.). PC3 cells (5×10⁶) were injected s.c. into 6-to 8-week-old male SCID mice, whereas MDA-MB-231 cells (2×10⁶) wereimplanted orthotopically into the mammary fat pad of female SCID mice.When the primary tumor volume reached approximately 150-200 mm³, animalswere randomized into eight groups (n=5-10/group). Each group was treatedwith either 0.9% saline/albumin vehicle control, Cremophor® EL vehiclecontrol, metronomic Taxol® (1.3 mg/kg, i.p., qd), metronomic ABI-007 (3,6, or 10 mg/kg paclitaxel, i.p., qd), MTD Taxol® (13 mg/kg, i.p., qd×5,1 cycle), or MTD ABI-007 (30 mg/kg paclitaxel, i.v., qd×5, 1 cycle).Perpendicular tumor diameters were measured with a caliper once a weekand their volumes were calculated. At the end of the treatment period,blood samples were drawn by cardiac puncture from mice in all groups.CECs and CEPs were enumerated as described herein.

Metronomic ABI-007 (3, 6 and 10 mg/kg) but not Taxol® (1.3 mg/kg)administered i.p. daily for 4 weeks significantly (p<0.05) inhibitedgrowth of both MDA-MB-231 and PC3 tumors (FIG. 3A and FIG. 3B). NeitherABI-007 nor Taxol® administered metronomically induced any weight loss(FIG. 3C and FIG. 3D). Although MTD ABI-007 (30 mg/kg) inhibited tumorgrowth more effectively than MTD Taxol® (13 mg/kg), significant weightloss was noted with the former, indicating toxicity. In addition, twoout of five mice treated with MTD ABI-007 displayed signs of paralysisin one limb 6 days after the last dose of drug. The paralysis wastransient and resolved within 24-48 hours. Interestingly, the antitumoreffect of metronomic ABI-007 at 6 mg/kg approximated that of MTD Taxol®in the MDA-MB-231, xenograft model (FIG. 3A). Increasing the dose ofmetronomic ABI-007 to 10 mg/kg did not seem to confer more pronouncedtumor growth inhibition. In contrast, metronomic ABI-007 elicitedgreater antitumor response at 10 mg/kg than at 3 and 6 mg/kg in the PC3xenografts (FIG. 3B).

Metronomic ABI-007 significantly decreased the levels of viable CEPs ina dose-dependent manner in MDA-MB-231 tumor-bearing mice (FIG. 4A).Viable CEP levels also exhibited a dose-dependent reduction in responseto metronomic ABI-007 in PC3 tumor-bearing mice, but reached statisticalsignificance only at 10 mg/kg (FIG. 4B). The levels of CEPs were notaltered by metronomic Taxol® in both xenograft models (FIGS. 4A and 4B).

Effects of metronomic and MTD ABI-007 and metronomic and MTD Taxol® onintratumoral microvessel density were studied. Five-um thick sectionsobtained from frozen MDA-MB-231 and PC3 tumors were stained with H&E forhistological examination by standard methods known to one skilled in theart. For detection of microvessels, sections were stained with a ratanti-mouse CD31/PECAM-1 antibody (1:1000, BD Pharmingen) followed by aTexas Red-conjugated goat anti-rat secondary antibody (1:200, JacksonImmunoResearch Laboratories, Inc., West Grove, Pa.). A singlemicrovessel was defined as a discrete cluster or single cell stainedpositive for CD31/PECAM-1d, and the presence of a lumen was not requiredfor scoring as a microvessel. The MVD for each tumor was expressed asthe average count of the three most densely stained fields identifiedwith a 20× objective on a Zeiss AxioVision 3.0 fluorescence microscopicimaging system. Four to five different tumors per each vehicle controlor treatment group were analyzed.

In MDA-MB-231 tumors, metronomic ABI-007 at 6 and 10 mg/kg as well asMTD ABI-007 seemed to reduce microvessel density (MVD) slightly althoughstatistical significance was not reached (FIG. 5A). In PC3 tumors,metronomic ABI-007 at 3 and 10 mg/kg appeared to decrease MVD butwithout reaching statistical significance (FIG. 5A). Interestingly, asignificant correlation existed between MVD and the level of viable CEPsin the MDA-MB-231 (FIG. 5B; r=0.76, P-0.04) but not in the PC3 (FIG. 5C;r=−0.071, P-0.88) model.

In vivo angiogenesis evaluation were carried out. A Matrigel plugperfusion assay was performed with minor modifications to methods knownby one skilled in the art. Briefly, 0.5 ml Matrigel supplemented with500 ng/ml of basic fibroblast growth factor (bFGF; R&D Systems Inc.,Minneapolis, Minn.) was injected s.c. on day 0 into the flanks of10-week-old female Balb/cJ mice. On day 3, animals were randomlyassigned to eight groups (n=5 each). Each group was treated with either0.9% saline/albumin vehicle control, Cremophor® EL vehicle control,metronomic Taxol® (1.3 mg/kg, i.p., qd), metronomic ABI-007 (3, 6, or 10mg/kg paclitaxel, i.p., qd), MTD Taxol® (13 mg/kg, i.v., qd×5), or MTDABI-007 (30 mg/kg paclitaxel, i.v, qd×5). As a negative control, fiveadditional female Balb/cJ mice of similar age were injected withMatrigel alone. On day 10, all animals were injected i.v. with 0.2 ml of25 mg/ml FITC-dextran (Sigma, St. Louis, Mo.). Plasma samples weresubsequently collected. Matrigel plugs were removed, incubated withDispase (Collaborative Biomedical Products, Bedford, Mass.) overnight at37° C., and then homogenized. Fluorescence readings were obtained usinga FL600 fluorescence plate reader (Biotech Instruments, Winooski, Vt.).Angiogenic response was expressed as the ratio of Matrigel plugfluorescence to plasma fluorescence.

Metronomic ABI-007 at 6 and 10 mg/kg appeared to decrease angiogenesisalthough the inhibition did not reach statistical significance (FIG. 6).Angiogenesis seemed to be unaltered by metronomic ABI-007 at 3 mg/kg,MTD ABI-007, MTD and metronomic Taxol® relative to the respectivevehicle controls (FIG. 6). These observations were similar to theintratumoral MVD results described herein.

Example 5 Nab-5109, A Nanoparticle Albumin-Bound IDN5109 (nab-5109)Shows Improved Efficacy and Lower Toxicity over the Tween® formulation(Tween®-5109, Ortataxel)

Methods: Nanoparticle nab-5109 was prepared using nab technology andcharacterized by laser light scattering. Nab-5109 and Tween-5109 weretested against Pgp+DLD-1 (known to be resistant against paclitaxel anddocetaxel—Vredenburg et al, JNCI 93: 1234-1245, 2001) human coloncarcinoma xenograft in nude mice (n=5/group) at doses of 50 mg/kg(Tween®-5109, previously shown as MTD) and 75 mg/kg (nab-5109) givenq3d×4, i.v. Control groups of PBS and human serum albumin (HSA) werealso used.

Results: Nab-5109 yielded nanoparticles with mean size, Z_(Ave)=119 nmand Zeta potential=−32.7 mV. Nab-5109 was lyophilized to a dry powderthat easily dispersed in saline. In vivo, there was significantly moreweight loss (ANOVA, p<0.001) in the tumor bearing animals withTween®-5109 (50 mg/kg, 8.8% wt loss) than with nab-5109 (75 mg/kg, 3.4%wt loss) indicating substantially lower toxicity of nab-5109 despite the50% higher dose. There was significant tumor suppression by nab-5109 andTween®-5109 (ANOVA, p<0.0001 vs. controls) with tumor growth delays of36 and 28 days respectively for nab-5109 (75 mg/kg) and Tween®-5109 (50mg/kg). Nab-5109 was more effective than Tween®-5109 (ANOVA, p=0.0001)in suppressing tumor growth. There were no differences between the PBSand HSA control group in term of toxicity and efficacy.

Conclusion: Nanoparticle albumin-bound, nab-5109 was successfullyprepared and could be given at 50% higher dose than Tween®-5109 withlower toxicity despite higher dose. At this higher dose, 75 mg/kg(q3d×4), nab-5109 showed significantly improved efficacy in thePgp+DLD-1 human colon xenograft compared with Tween®-5109.

Example 6 Nanoparticle Albumin Bound (nab) Dimeric Thiocolchicinesnab-5404, nab-5800, and nab-5801: A Comparative Evaluation of AntitumorActivity vs Abraxane™ and Irinotecan

Methods: Nanoparticle colchicines were prepared using nab technology.Cytotoxicity was evaluated in vitro using human MX-1 breast carcinomacultures. In vivo anti-tumor activity (human HT29 colon tumor xenograft)in nude mice was compared against Irinotecan and Abraxane™. Dose levelsfor the nab-colchicines and Irinotecan were 20 mg/kg, 30 mg/kg, and 40mg/kg, given q3d×4, i.v. Abraxane™ was dosed at its MTD, 30 mg/kg, givenqd×5.

Results: The hydrophobic thiocolchicine dimers yielded nanoparticleswith average size Z_(Ave)(nm) of 119, 93, and 84 for nab-5404, nab-5800,and nab-5801, respectively. The nanoparticle suspensions were sterilizedthrough 0.22 um filters and lyophilized. In vitro, nab-5404 was the mostpotent of the three analogs against MX-1 (p≦0.0005, ANOVA), (IC₅₀(ug/ml): 18, 36 and 77 for nab-5404, nab-5800 and nab-5801 respectively)as well as against the HT29 xenograft in vivo (p<0.0001, ANOVA). Tumorvolume was suppressed by 93%, 79%, and 48% with nab-5404 at doses 40mg/kg, 30 mg/kg, and 20 mg/kg, respectively. In contrast, tumor volumewas only suppressed by 31%, 16%, and 21% with nab-5800; and 17%, 30%,and 23% with nab-5801 at 40 mg/kg, 30 mg/kg, and 20 mg/kg, respectively.Nab-5404 was more effective than Irinotecan at all dose levels (p≦0.008,ANOVA) with tumor volumes for Irinotecan suppressed by only 48%, 34%,and 29% at dose levels of 40 mg/kg, 30 mg/kg, and 20 mg/kg,respectively. In comparison to Abraxane™, nab-5404 was more active atequitoxic dose (ETD) based on equal weight loss (p<0.0001, ANOVA). Tumorvolume was suppressed 93% by nab-5404 (40 mg/kg, q4d×3) and 80% byAbraxane™ (30 mg/kg, qd×5) at their respective ETDs.

Conclusions: Nab technology was utilized to convert 3 hydrophobicdimeric thiocolchicines (IDN5404, IDN5800, IDN5801) to nanoparticlessuitable for I.V. administration. Nab-5404 had superior antitumoractivity in vitro and in vivo compared to nab-5800 and nab-5801.Nab-5404 was more potent than Irinotecan at equal dose. At equitoxicdose, defined by weight loss, nab-5404 was more potent than Abraxane™.These data warrant further investigation of nab-5404.

Example 7 Abraxane™ vs Taxotere®: A Preclinical Comparison of Toxicityand Efficacy

Methods: Toxicity of Abraxane™ and Taxotere® was compared in adose-ranging study in nude mice given the drugs on a q4d×3 schedule. Thedose levels were Taxotere® 7, 15, 22, 33, and 50 mg/kg and ABX 15, 30,60, 120, and 240 mg/kg. Antitumor activity of Abraxane™ and Taxotere®was compared in nude mice with human MX-1 mammary xenografts at a doseof 15 mg/kg weekly for 3 weeks.

Results: In the dose-escalation study in mice, the Taxotere® maximumtolerated dose (MTD) was 15 mg/kg and lethal dose (LD₁₀₀) was 50 mg/kg.In contrast, the Abraxane™ MTD was between 120 and 240 mg/kg and LD₁₀₀was 240 mg/kg. In the tumor study Abraxane™ was more effective thanequal doses of Taxotere® in tumor growth inhibition (79.8% vs 29.1%,p<0.0001, ANOVA).

Conclusion: Nanoparticle abumin-bound paclitaxel (Abraxane™) wassuperior to Taxotere® in the MX-1 tumor model when tested at equaldoses. Furthermore, the toxicity of Abraxane™ was significantly lowerthan that of Taxotere®, which would allow dosing of Abraxane atsubstantially higher levels. These results are similar to the enhancedtherapeutic index seen with Abraxane™ compared to Taxol® and suggestthat the presence of surfactants may impair the transport, antitumoractivity and increase the toxicity of taxanes. Studies in additionaltumor models comparing Abraxane™ and Taxotere® are ongoing.

Example 8 A Nanoparticle Albumin Bound Thiocolchicine dimer (nab-5404)with Dual Mechanisms of Action on Tubulin and Topoisomerase-1:Evaluation of In vitro and In vivo Activity

Methods: IDN5404 was tested for cytotoxic activity using the MCF7-Sbreast carcinoma and its multidrug resistant variant, MCF7-R (pgp+). Itscytotoxicity was also assessed against the NCI-60 human tumor cell linepanel. The nanoparticle albumin bound nab-5404 was administered IV usingvarious schedules, to SCID mice implanted s.c. with a human A121 ovariantumor xenograft.

Results: Against MCF7 cell lines, the parent compound, colchicine,demonstrated tumor growth inhibition with the IC50 value (50% growthinhibitory concentration) for MCF7-S cells at 3.9±0.2 nM. The resistantvariant MCF7-R demonstrated an IC50 of 66±8.6 nM, approximately a17-fold increase due to drug resistance. IDN5404, demonstrated increasedactivity against both cell lines, displaying IC50 values of 1.7±0.1 and40±3.8 nM, respectively. These results were confirmed within the NCI 60human tumor cell line panel with IDN5404 having a mean IC50 of <10⁻⁸Mand >10 fold resistance between the MCF7-S and the MCF7-R cell lines.The COMPARE algorithm identified IDN5404 as a tubulin binder similar tovinca alkaloids, confirming the previous results. In vivo against theA121 ovarian tumor xenograft, efficacy and toxicity of nab-5404 was doseand schedule dependent. Nanoparticle nab-5404 was well tolerated andcapable of inducing complete regressions and cures: at 24 mg/kgadministered IV qd×5, 5 of 5 mice were long-term survivors (LTS) with noevidence of tumor. However, increasing the dosage to 30 mg/kg resultedin 5 of 5 toxic deaths. On a q3d×4 schedule, 30 mg/kg resulted in 4 of 5mice LTS and at 50 mg/kg, 5 of 5 toxic deaths. Using a q7d×3 schedule,40 mg/kg resulted in 3 of 5 mice LTS and at 50 mg/kg, 4 of 4 LTS werenoted.

Conclusions: IDN5404, a new thiocolchicine dimer with dual mechanism ofaction showed activity in pgp-expressing, cisplatin and topotecanresistant cell lines. In vivo, nanoparticle albumin bound nab-5404 wasactive against A121 ovarian xenografts.

Example 9 Combination Studies of Abraxane™ and Other Agents

Due to the advantageous properties of Abraxane™ (ABX, the nanoparticlealbumin-bound paclitaxel) noted above, it was used and being used in anumber of studies with different modes of administration and schedulesand in combination with other oncology drugs as well as radiationtreatment. These are listed below:

In metastatic breast cancer, these studies include: Randomized Phase IITrial of ABX 125, Gem 1000 mg/m², To evaluate the combination of ABXWeekly Abraxane ™ in Combination D1,8; q 3wk and Gemcitabine in 1st-lineMBC. with Gemcitabine in Individuals with HER2 Negative MetastaticBreast Cancer A phase II study of weekly ABX 100 mg/m², Carbo AUC Datawill be important for using dose-dense nanoparticle paclitaxel 2, bothD1,8,15; Her 2 mg/kg ABX in combination with carbo (ABI-007)carboplatin, with (4 mg/kg on wk a) q4wk × 6 and/or Herceptin ®. Alsohelpful for Herceptin ® as first or second-line other combinations.therapy of advanced HER2 positive breast cancer Weekly Vinorelbine andL1: ABX 80, Nav 15; L2: Multi-center study of ABX in Abraxane ™, with orwithout G-CSF, ABX 90, Nav 20; L3: ABX combination with Navelbine ® inin stage IV breast cancer: a phase 100, Nav 22.5; L4: ABX 110, 1st-lineMBC. I-II study Nav 25; L5: ABX 125, Nav 25 qwk Phase II trial of weeklyAbraxane ™ ABX 125 mg/m² Q¾wk A relatively large phase II of weeklymonotherapy for 1st-line MBC (plus ABX monotherapy at 125 mg/m² inHerceptin ® in Her2+ pts) 1st-line MBC. Phase I/II trial Abraxane ™ plusABX + Anthracycline Doxil ® for MBC plus limited PK 3-arm phase II trialin 1st-line MBC ABX weekly (130 mg/m²) vs. To optimize ABX monotherapyq2wk (260 mg/m²) vs. q3wk regime for MBC (260 mg/m²) 3-arm phase IItrial in 1st-line and ABX weekly vs. ABX q3wk randomized ABX MBC trialto 2nd-line MBC, with biological vs. Taxol ® weekly obtain importantdata: weekly ABX correlates analyses vs. weekly Taxol ®; weekly ABX vs.3-weekly ABX; plus biomarker study (caveolin-1 and SPARC). Phase I/IIAbraxane ™ + GW572016 TBD combination of ABX and GW572016 (a dual EGFRinhibitor and one of the most promising new biological agents for BC). Aphase I dose escalation study of a Abraxane ™ 100 mg/m² This phase Itrial is to determine the 2 day oral gefitinib weekly, 3 out of 4 weeks;safety and tolerability of a 2 day chemosensitization pulse given priorGefitinib starting at 1000 mg/d × 2 gefitinib pulse given prior to toweekly Abraxane ™ in individuals days Abraxane ™ administration. withadvanced solid tumors Phase II 1^(st) line MBC trial weekly ABX (125mg/m², 2 wk To evaluate the combination of ABX on and 1 wk off) +Xeloda ® and Xeloda ® in 1st-line MBC, using 825 mg/m² d 1-14 q3wk 2weekly on and 1 weekly off ABX regime. Phase II pilot adjuvant trial ofDose dense AC + G CSF --> A pilot adjuvant study of a “super Abraxane ™in breast cancer weekly ABX --> Avastin ® dose dense” Abraxane ™ indose-dense adjuvant AC q2w × 4 + G CSF --> ABX A pilot adjuvant study ofdose dense chemotherapy for early stage breast q2wk × 4 ABX regime - analternate of a cancer standard adjuvant regime Phase II pilot adjuvanttrial of AC Q2wk --> ABX q2wk + G- A pilot adjuvant study in preparationAbraxane ™ in breast cancer CSF for phase III adjuvant trial

In Breast cancer neodajuvant setting studies include: Phase II Trial ofDose Dense Neoadjuvant: Gem 2000, This neoadjuvant study is based on theNeoadjuvant Gemcitabine, Epirubicin, Epi 60, ABX 175 mg/m², GET datafrom Europe which showed ABI-007 (GEA) in Locally Advanced Neul 6 mg SC,all D1 q2 high activity. In the current regime, or Inflammatory BreastCancer. wk × 6 Adjuvant: Gem ABX will replace T, or Taxol ®. 2000, ABX220, Neul 6 mg D1 q2wk × 4 Phase II preoperative trial of ABX 220 mg/m²q2wk × 6 Abraxane ™ followed by FEC (+Herceptin ® followed by FEC × 4 asappropriate) in breast (+Herceptin ® for Her2+ cancer pts) Pre-clinicalstudy of drug-drug ABX + other agents interaction Phase II neoadjuvant(ABX + Herceptin ®) followed by (Navelbine ® + Herceptin ®) Randomizedphase II trial of TAC vs. AC followed To evaluate AC followed byneoadjuvant chemotherapy in ABX + carbo vs. AC ABX/carbo orABX/carbo/Herceptin ® individuals with breast cancer followedcombinations vs TAC (a FDA ABX + carbo + Herceptin ® approved adjuvantBC regime) in neoadjuvant setting. Phase II neoadjuvant trial of ABX:200 mg/m² D1; Abraxane ™ and capecitabine in Xel: 1000 mg/m² D1-14;locally advanced breast cancer q3wk × 4 Phase II trial of neoadjuvantABX: 300 mg/m² q3wk chemotherapy (NCT) with nanoparticle paclitaxel(ABI-007, Abraxane ™) in women with clinical stage IIA, IIB, IIIA, IIIB,and IV (with intact primary) breast cancers

In lung cancer the studies include: Phase I/II study of Abraxane ™ ABXweekly The first phase II trial of ABX monotherapy in 1st-line advancedcombo with carbo in NSCLC. NSCLC Phase II Trial of weekly Abraxane ™ABX: 125 mg/m² plus carboplatin in 1st-line NSCLC D1,8,15; Carbo: AUC 6D1; q4 wk A Phase I Trial of Carboplatin and Arm 1: ABX 100, 125, This2-arm phase I study will Abraxane ™ on a weekly and every 150 mg/m²D1,8,15 generate important data on three week schedule in individualsq4wk; Arm 2: ABX ABX/carbo combination for with Advanced Solid Tumor220, 260, 300 mg/m² further studies of this combo Malignancies D1 q3wk.Carbo in multiple diseases. AUC6 in both arms Phase II study of ABI 007ABX Level(a): 225 mg/m2; This phase II NSCLC study (Abraxane ™) andcarboplatin in Level(b): 260 mg/m2; will generate data for a futureadvanced non-small cell lung cancer. Level(3): 300 mg/m2; phase IIIregistration trial in q3wk Carbo lung cancer fixed at AUC6 q3wk Phase Istudy of ABI 007 ABX q3wk (Abraxane ™) and carboplatin Phase I/II studyof Abraxane ™ + Alimta ® TBD ABX and Alimta ® can be a for 2nd-lineNSCLC promising combination due to the non-overlapping toxicityprofiles. Phase I/II trial of Abraxane ™ plus cisplatin in advancedNSCLC Phase I/II study of Abraxane ™, Navelbine ®, and Cisplatin fortreatment of advanced NSCLC Phase II ABX mono in 1st-line ABX 260 mg/m²q3wk The 1st ABX trial in NSCLC. NSCLC Phase II study of Abraxane ™Cohort 1: ABX q3wk; monotherapy in 2nd-line NSCLC Cohort 2: ABX weekly.Doses TBD Phase I/II trial of weekly Abraxane ™ 1st line and carboplatinin advanced NSCLC

Studies in Prostate include: Randomized phase II ABX 100 mg/m² weekly vsQ3W in front line weekly vs 260 mg/m² HRP q3wk Phase II ABX in 1st-lineweekly ABX Phase II study of weekly ABX in 1st-line prostate cancer HRPCPhase II neoadjuvant study TBD A multi-center neoadjuvant trial of ABXin prostate cancer plus biomarker study. Phase II ABX 100 mg weekly nobreak

Studies in ovarian cancer include: Phase II study of Abraxane ™ TBD fortreatment of advanced ovarian cancer (3rd-line) Phase I study ofAbraxane ™ ABX weekly + Carbo plus carbo for treatment of AUC 6 advancedovarian cancer A phase II trial of Abraxane ™/Carboplatin in recurrentovarian cancer

Studies in Chemoradiation include: Phase I/II trial of Abraxane ™combined with radiation in NSCLC Abraxane ™ Combined With animal modelRadiation H&N (Head and Neck Cancer) TBD

Other studies include: Phase II study of ABX in treatment of 125 mg/m²d1, 8, 15 persistent or recurrent carcinoma of the q28 days cervix PhIIin preciously treated (100 ABX) 26-->70 and untreated (150 ABX)metastatic melanoma Ph II single treatment use of ABI-007 for thetreatment of non-hematologic malignancies Abraxane ™ Combined Withantiangiogenic agents, e.g., Avastin ®. Abraxane ™ Combined Withproteasome inhibitors e.g., Velcade ®. Abraxane ™ Combined With EGFRinhibitors e.g., Tarceva ®. A randomized phase II trial of weeklygemcitabine, Abraxane ™, and external irradiation for locally advancedpancreatic cancer

Example 10 Combination of Nanoparticle Invention Drugs with Other Agentsand Modes of Therapy

Lower toxicity of nanoparticle invention drugs described herein allowcombination with other oncology drugs and other modes of treatment withmore advantageous outcome. These include nanoparticle forms ofpaclitaxel, docetaxel, other taxanes and analogs, geldanamycins,colchicines and analogs, combretastatins and analogs, hydrophobicpyrimidine compounds, lomaiviticins and analogs including compounds withthe lomaiviticin core structures, epothilones and analogs,discodermolide and analogs and the like. The invention drugs may becombined with paclitaxel, docetaxel, carboplatin, cisplatin, otherplatins, doxorubicin, epirubicin, cyclophosphamide, iphosphamide,gemcitabine, capecitabine, vinorelbine, topotecan, irinotecan,tamoxifen, camptothecins, 5-FU, EMP, etoposide, methotraxate and thelike.

Example 11 Combination of Abraxane™ with Carboplatin and Herceptin®

The combination of Taxol® and carboplatin has shown significant efficacyagainst metastatic breast cancer. On a weekly schedule, in thiscombination, Taxol® can only be dosed at up to 80 mg/m². Higher dosescannot be tolerated due to toxicity. In addition, HER-2-positiveindividuals derive greater benefit when Herceptin® is included in theirtherapeutic regime. This open-label Phase II study was conducted todetermine the synergistic therapeutic effect of ABI-007 (Abraxane™) withthese agents. The current study was initiated to evaluate the safety andantitumor activity of ABI-007/carboplatin with Herceptin® forindividuals with HER-2 positive disease. ABI-007 was given incombination with carboplatin and Herceptin® administered intravenouslyweekly to individuals with HER-2 positive advanced breast cancer. Acohort of 3 individuals received ABI-007 at a dose of 75 mg/m² IVfollowed by carboplatin at target AUC=2 weekly and Herceptin® infusion(4 mg/kg at week 1, and 2 mg/kg on all subsequent weeks) for 1 cycle.These individuals tolerated the drug very well so for all subsequentcycles and individuals the dose of ABI-007 was escalated to 100 mg/m².Six individuals were treated to date. Of the 4 individuals that wereevaluated for response, all 4 (100%) showed a response to the therapy.It should be noted that due to lower toxicity of Abraxane™, a highertotal paclitaxel dose could be given compared to Taxol® with resultingbenefits to the individuals.

Example 12 Combination of Abraxane™ with Carboplatin

The combination of Taxol® and carboplatin has shown significant efficacyin lung cancer. Another study with Abraxane™ in combination withcarboplatin on a 3 weekly schedule in individuals with lung cancer isongoing.

Example 13 Use of Abraxane™ in Combination With Radiation Example 13a

Abraxane™, combined with clinical radiotherapy, enhances therapeuticefficacy and reduces normal tissue toxicity. Abraxane™ is used toincrease the therapeutic gain of radiotherapy for tumors; to enhancetumor response to single and fractionated irradiation; to enhance normaltissue response to radiation and to increase therapeutic ratio ofradiotherapy.

A murine ovarian carcinoma, designated OCa-I, which has beeninvestigated extensively is used. First, optimal timing of Abraxane™administration relative to local tumor radiation is timed to producemaximum antitumor efficacy. Tumors are generated in the right hind legof mice by i.m. injection of tumor cells and treatment is initiated whenthe tumors reach. 8mm in size. Mice are treated with 10 Gy single doseirradiation, a single dose of Abraxane™ or with combination therapy ofAbraxane™ given at different times 5 days before to 1 day afterirradiation. A dose of Abraxane™ equal to about 1½ times more than themaximum tolerated dose of paclitaxel is used, a dose of 90 mg/kg. Theendpoint of efficacy is tumor growth delay. The groups consist of 8 miceeach. Tumors are generated and treated-as described in Aim 1. Theendpoint of efficacy is tumor growth delay. Tumors are irradiated with5, 7.5 or 10 Gy delivered either in a single dose or in fractionateddoses of 1, 1.5 or 2 Gy radiation daily for five consecutive days. SinceAbraxane™ is retained in the tumor for several days and exerts itsenhancing effect on each of the five daily fractions, Abraxane™ is givenonce at the beginning of the radiation regime. Since the ultimate goalin clinical radiotherapy is to achieve tumor cure, the potential forAbraxane™ to enhance tumor radiocurability is determined. The samescheme as described for the fractionated tumor growth delay study isused, except that a range of doses from 2 to 16 Gy is given daily forfive consecutive days (total radiation dose 10 to 80 Gy). Tumors arefollowed for regression and regrowth for up to 120 days afterirradiation, when TCD50 (the dose of radiation needed to yield localtumor cure in 50 percent of animals) is determined. There are two TCD50assays: radiation only and Abraxane™ plus radiation, and each assayconsists of 10 radiation dose groups containing 15 mice each. To providetherapeutic gain, any radioenhancing agent, including Abraxane™, mustincrease tumor radioresponse more than increase normal tissue damage byradiation. Damage to jejunal mucosa, a highly proliferative tissueaffected by taxanes is assessed. The jejunal microcolony assay is usedto determine the survival of crypt epithelial cells in the jejunum ofmice exposed to radiation. Mice are exposed to whole body irradiation(WBI) with daily doses of X-rays ranging from 3 to 7 Gy for fiveconsecutive days. The mice are treated with Abraxane™, at an equivalentpaclitaxel dose of 80 mg/kg, administered i.v. 24 h before the firstdose of WBI and killed 3.5 days after the last dose of WBI. The jejunumis prepared for histological examination, and the number of regeneratingcrypts in the jejunal cross-section is counted. To construct radiationsurvival curves, the number of regenerating crypts is converted to thenumber of surviving cells.

Example 13b

The objective of this study was to assess whether ABI-007 (a) as asingle agent has antitumor activity against the syngeneic murine ovariancarcinoma OCa-1 and (b) enhances the radiation response of OCa-1 tumorsin a combined treatment regime as described in the previous example withthe following modifications.

OCa-1 tumor cells were injected i.m. into the hind leg of C3H mice. Whentumors grew to a mean diameter of 7 mm, single treatment with localradiation (10 Gy) to the tumor-bearing leg, ABI-007 90 mg/kg i.v., orboth, was initiated. To determine the optimal treatment scheduling,ABI-007 was given from 5 days to 9 hours before radiation as well as 24hours after radiation. Treatment endpoint was absolute tumor growthdelay (AGD), defined as the difference in days to grow from 7-12 mm indiameter between treated and untreated tumors. For groups treated withthe combination of ABI-007 and radiation, an enhancement factor (EF) wascalculated as the ratio of the difference in days to grow from 7 to 12mm between the tumors treated with the combination and those treatedwith ABI-007 alone to the AGD of tumors treated with radiation only. Toassess the radiation-enhancing effect of ABI-007 for a fractionatedradiation regime on the endpoint tumor cure, a TCD50 assay was performedand analyzed 140 days post treatment. Total doses of 5 to 80 Gy in 5daily fractions were administered either alone or combined with ABI-00724 hours before the first radiation dose.

As a single agent, ABI-007 significantly prolonged the growth delay ofthe OCa-1 tumor (37 days) compared to 16 days for untreated tumors.ABI-007 as a single agent was more effective than a single dose of 10Gy, which resulted in a delay of 29 days. For combined treatmentregimes, ABI-007 given at any time up to 5 days before radiation,produced a supra-additive antitumor effect. EF was 1.3, 1.4, 2.4, 2.3,1.9, and 1.6 at intertreatment intervals of 9 h, 24 h and 2, 3, 4, and 5days, respectively. When ABI-007 was given after radiation, the combinedantitumor treatment effect was less than additive. Combined treatmentwith ABI-007 and radiation also had a significant effect on tumor cureby shifting the TCD50 of 55.3 Gy for tumors treated with radiation onlyto 43.9 Gy for those treated with the combination (EF 1.3).

This experiment demonstrated that ABI-007 possesses single-agentantitumor activity against OCa-1 and enhances the effect of radiotherapywhen given several days prior. As previously demonstrated for paclitaxeland docetaxel, the radiation enhancement is likely a result of multiplemechanisms, with a cell cycle arrest in G2/M being dominant at shorttreatment intervals and tumor reoxygenation at longer intervals.

Example 14 Combination of Abraxane™ and Tyrosine Kinase Inhibitors

Pulse-dosing of gefitinib in combination with the use of Abraxane™ isuseful to inhibit the proliferation of EGFr expressing tumors. 120 nudemice are inoculated with BT474 tumor cells to obtain at least 90 micebearing BT474 xenograft tumors and split into 18 experimental arms (5mice each). Arm 1 mice receive control i.v. injections. All other micereceive weekly i.v. injections of Abraxane™ at 50 mg/kg for 3 weeks. Arm2 receive Abraxane™ alone. Arms 3, 4, 5, 6, 7, 8 receive weeklyAbraxane™ preceded by 2 days of a gefitinib pulse at increasing doses.Arms 9, 10, 11, 12, 13 receive weekly Abraxane™ preceded by one day of agefitinib pulse at increasing doses. Arms 14, 15, 16, 17, 18 receiveweekly Abraxane™ along with everyday administration of gefitinib atincreasing doses. The maximum tolerated dose of gefitinib that can begiven in a 1 or 2 day pulse preceding weekly Abraxane™ or in continuousadministration with Abraxane™ is established. In addition, measurementof anti-tumor responses will determine whether a dose-responserelationship exists and whether 2 day pulsing or 1 day pulsing issuperior. These data are used to select the optimal dose of pulsegefitinib and that of continuous daily gefitinib given with Abraxane™.

120 nude mice are inoculated with BT474 tumor cells to obtain 90 micebearing tumors. These mice are split into 6 groups (15 each). Arm 1receive control i.v. injections. Arm 2 receive Abraxane™ 50 mg/kg i.v.weekly for 3 weeks. Arm 3 receive oral gefitinib at 150 mg/kg/day. Arm 4receive Abraxane TM50 mg/kg along with daily gefitinib at the previouslyestablished dose. Arm 5 receive Abraxane™ 50 mg/kg preceded by agefitinib pulse at the previously established dose and duration. Arm 6receive only a weekly gefitinib pulse at the previously establisheddose. After three weeks of therapy, mice are followed until controlsreach maximum allowed tumor sizes.

Example 15. Phase II Study of Weekly, Dose-Dense Nab™-Paclitaxel(Abraxane™) Carboplatin With Trastuzumab® As First-line Therapy OfAdvanced HER-2 Positive Breast Cancer

This study aimed to evaluate (1) the safety and tolerability and (2) theobjective response rate of weekly dose-densetrastuzumab/Abraxane™/carboplatin as first-line cytotoxic therapy forpatients with advanced/metastatic (Stage IV adenocarcinoma)HER-2-overexpressing breast cancer. Trastuzumab is a monoclonalantibody, also known as Herceptin®, which binds to the extracellularsegment of the erbB2 receptor.

Briefly, patients without recent cytotoxic or radiotherapy wereincluded. Doses of Abraxane™ were escalated from 75 mg/m² as 30-min i.v.infusions on days 1, 8, 15 up to 100 mg/m² for subsequent cyclesaccording to the standard 3+3 rule. Carboplatin AUC=2 was given as 30-60min i.v. infusions on days 1, 8, 15 and for an initial 29 day cycle.Trastuzumab was given as i.v. 30-90 min infusion on days 1, 8, 15, 22 ata dose of 4 mg/kg at week 1 and 2 mg/kg on all subsequent weeks.

Of 8 out of 9 patients evaluable for response the response rate(confirmed plus unconfirmed) was 63% with 38% stable disease. The mostcommon toxicities were neutropenia (grade 3: 44%; grade 4: 11%) andleukocytopenia (33%).

These results suggest that trastuzumab plus Abraxane™ plus carboplatindemonstrated a high degree of antitumor activity with acceptabletolerability as a first-line therapy for MBC.

Example 16 Phase II Trial of Capecitabine Plus nab™-Paclitaxel(Abraxane™) in the First Line Treatment of Metastatic Breast Cancer

The purpose of this phase II study was to evaluate the safety, efficacy(time to progression and overall survival), and quality of life ofpatients with MBC who received capecitabine in combination withAbraxane™. Capecitabine is a fluoropyrimidine carbamate also known asXeloda® which has been shown to have substantial efficacy alone and incombination with taxanes in the treatment of MBC.

In this open-label, single-arm study, Abraxane™ 125 mg/m² was given byi.v. infusion on day 1 and day 8 every 3 weeks plus capecitabine 825mg/m² given orally twice daily on days 1 to 14 every 3 weeks. Patientswere HER-2/neu negative with a life expectancy of greater than 3 months.Patients had no prior chemotherapy for metastatic disease, no priorcapecitabine therapy, and no prior fluoropyrimidine therapy andpaclitaxel chemotherapy given in an adjuvant setting.

12 patients have been enrolled with safety analysis completed on thefirst 6 patients and the response rate evaluable after 2 cycles in thefirst 8 patients. There were no unique or unexpected toxicities with nograde 4 toxicities or neuropathy greater than grade 1. Response datawere confirmed on only the first 2 cycles of therapy (first evaluationpoint) in 6 patients. Two patients have completed 6 cycles with 1partial response and 1 stable disease. Of the first 8 patients after 2cycles, there were 2 partial responses and 4 with stable disease.

These results show that combination of capecitabine and weekly Abraxane™at effective doses is feasible with no novel toxicities to date.Abraxane™ related toxicity was mainly neutropenia without clinicalconsequences, and hand foot syndrome was the major toxicity ofcapecitabine.

Example 17 Pilot Study of Dose-Dense Doxorubicin Plus CyclophosphamideFollowed by nab-paclitaxel (Abraxane™) in Patients with Early-StageBreast Cancer

The objective of this study was to evaluate the toxicity of doxorubicin(adriamycin) plus cyclophosphamide followed by Abraxane™ in early stagebreast cancer.

Patients had operable, histologically confirmed breast adenocarcinoma ofan early stage. The patients received doxorubicin (adriamycin) 60 mg/m²plus cyclophosphamide 600 mg/mm² (AC) every 2 weeks for 4 cyclesfollowed by Abraxane™ 260 mg/m² every two weeks for 4 cycles.

30 patients received 4 cycles of AC, and 27 of 29 patients received 4cycles of Abraxane™; 33% of patients received pegfilgrastim (Neulasta®)for lack of recovery of ANC (absolute neutrophil count) duringAbraxane™. Nine patients (31%) had Abraxane™ dose reductions due tonon-hematologic toxicity. A total of 9 patients had grade 2 and 4patients had grade 3 peripheral neuropathy (PN); PN improved by ≧1 gradewithin a median of 28 days.

These results indicate that dose-dense therapy with doxorubicin (60mg/m²) plus cyclophosphamide (600 mg/m²) every 2 weeks for 4 cyclesfollowed by dose-dense Abraxane™ (260 mg/m²) every 2 weeks for 4 cycleswas well tolerated in patients with early-stage breast cancer.

Example 18 Weekly nab-Paclitaxel (Abraxane™) as First Line Treatment ofMetastatic Breast Cancer with Trastuzumab Add On for HER-2/neu-PositivePatients

The purpose of the current study was to move weekly Abraxane to afront-line setting and add trastuzumab for HER2/neu-positive patients.

This phase II, open-label study included 20 HER2-postivive and 50HER2-negative patients with locally advanced or metastatic breastcancer. Abraxane™ was given at 125 mg/m² by 30 minute i.v. infusion ondays 1, 8, and 15 followed by a week of rest. Trastuzumab was givenconcurrently with study treatment for patients who were HER2-positive.The primary endpoint was response rate and the secondary endpoints weretime to progression (TTP), overall survival (OS), and toxicity.

In the safety population, 23 patients received a median of 3 cycles ofAbraxane™ to date. The most common treatment-related adverse event wasgrade 3 neutropenia (8.7%) with no grade 4 adverse events. One out of 4evaluable patients responded-to therapy.

Example 19 Phase I Trial of nab-Paclitaxel (Abraxane™) and Carboplatin

The aim of the current study was to determine the maximum tolerated doseof Abraxane™ (both weekly and every 3 weeks) with carboplatin AUC=6 andto compare the effects of sequence of administration on pharmacokinetics(PK).

Patients with histologically or cytologically documented malignancy thatprogressed after “standard therapy” were included. Arm 1 receivedAbraxane™ every 3 weeks in a dose escalation format based on cycle 1toxicities (220, 260, 300, 340 mg/m²) every 3 weeks followed bycarboplatin AUC=6. Arm 2 received weekly (days 1, 8, 15 followed by 1week off) Abraxane™ (100, 125, 150 mg/m²) followed by carboplatin AUC=6.For the PK portion of the study, Abraxane™ was followed by carboplatinin cycle 1 and the order of administration reversed in cycle 2 with PKlevels determined at initial 6, 24, 48 and 72 hours.

On the every 3 weeks schedule, neutropenia, thrombocytopenia andneuropathy were the most common grade 3/4 toxicities (3/17 each). On theweekly schedule, neutropenia 5/13 was the most common grade 3/4toxicity. The best responses to weekly administration at the highestdose of 125 mg/² (n=6) were 2 partial responses (pancreatic cancer,melanoma) and 2 stable disease (NSCLC). The best responses to the everythree week administration at the highest dose of 340 mg/m² (n=5) were 1stable disease (NSCLC) and 2 partial responses (SCLC, esophageal).

These data indicate activity of combination of Abraxane™ andcarboplatin. The MTD for the weekly administration was 300 mg/m², andfor the once every 3 week administration was 100 mg/m².

Example 20 Phase II Trial of Dose-Dense Gemcitabine, Epirubicin, andnab-Paclitaxel (Abraxane™) (GEA) in Locally Advanced/Inflammatory BreastCancer

In an open-label, phase II study an induction/neoadjuvant therapy regimewas instituted prior to local intervention. The therapy regime wasgemcitabine 2000 mg/m² i.v. every 2 weeks for 6 cycles, epirubicin 50mg/m² every 2 weeks for 6 cycles, Abraxane™ 175 mg/m² every 2 weeks for6 cycles, with pegfilgrastim 6 mg s.c. on day 2 every 2 weeks. Thepostoperative/adjuvant therapy regime after local intervention wasgemcitabine 2000 mg/m² every 2 weeks for 4 cycles, Abraxane™ 220 mg/m²every 2 weeks for 4 cycles and pegfilgrastim 6 mg s.c. day every 2weeks. Patients included females with histologically confirmed locallyadvanced/inflammatory adenocarcinoma of the breast.

Example 21 Cytotoxic Activity of Nab-Rapamycin in Combination withAbraxane™ on Vascular Smooth Muscle Cells

Vascular smooth muscle cells (VSMC) were seeded onto 96 wells plates inthe presence of increasing concentrations of nab-rapamycin and 0 μM, 1μM, 10 μM, or 100 μM of Abraxane™ (ABI-007). To evaluate the cytotoxiceffect of nab-rapamycin and Abraxane™, treated VSMCs were stained withethidium homodimer-1 (Invitrogen, Carlsbad Calif.) and analyzed for redfluorescence. Ethidium homodimer-1 is a high-affinity, fluorescentnucleic acid stain that is only able to pass through compromisedmembranes of dead cells to stain nucleic acids. As shown in FIG. 7A,nab-rapamycin, by itself, exhibited dose-dependent cell killing asdemonstrated by increasing fluorescence. Cell killing by nab-rapamycinwas not enhanced by Abraxane™ at 1 μM or 10 μM; however, it was greatlyenhanced by Abraxane™ at 100 μM (ANOVA, p<0.0001). Cells stained withethidium homodimer-1 as shown in FIG. 7A were also exposed to calcein.Calcein AM (Invitrogen) is a non-fluorescent molecule that is hydrolyzedinto fluorescent calcein by nonspecific cytosolic esterases. Live cellsexposed to calcein AM exhibit bright green fluorescence as they are ableto generate the fluorescent product and retain it. As shown in FIG. 7B,nab-rapamycin exhibited dose dependent cytotoxic activity as shown by areduced amount of fluorescent staining by calcein. This reduction influorescence was enhanced by coincubation with Abraxane™ in a dosedependent manner. ANOVA statistic gave p<0.0001 at all drugconcentrations of Abraxane™.

Example 22 Cytotoxic Activity of Nab-Rapamycin in Combination withAbraxane™ Against HT29 (Human Colon Carcinoma) Tumor Xenograft

Nude mice were implanted with 106 HT29 cells on their right flanks.Treatment was initiated once the tumor were palpable and were greaterthan 100-200 mm³. The mice were randomly sorted into 4 groups (n=8 pergroup). Group 1 received saline 3 times weekly for 4 weeks, i.v.; Group2 received Abraxane™ at 10 mg/kg, daily for 5 days, i.p.; Group 3received nab-rapamycin at 40 mg/kg, 3 times weekly for 4 weeks, i.v.;and Group 4 received both nab-rapamycin (40 mg/kg, 3 times weekly for 4weeks, i.v.) and Abraxane™ (10 mg/kg, daily for 5 days, i.p.). As shownin FIG. 8, the tumor suppression was greater for the Abraxane™ plusnab-rapamycin combination therapy than for either single therapy group.

Example 23 Cytotoxic Activity of Nab-17-AAG in Combination withAbraxane™ Against H358 (Human Lung Carcinoma) Tumor Xenograft

Nude mice were implanted with 10⁷ H358 cells on their right flanks.Treatment was initiated once the tumors were palpable and were greaterthan 100-200 mm³. The mice were randomly sorted into 4 groups (n=8 pergroup). Group 1 received saline 3 times weekly for 4 weeks, i.v.; Group2 received Abraxane™ at 10 mg/kg, daily for 5 days, i.p.; Group 3received nab-17-AAG at 80 mg/kg, 3 times weekly for 4 weeks, i.v.; andGroup 4 received both nab-17-AAG (80 mg/kg, 3 times weekly for 4 weeks,i.v.) and Abraxane™ (10 mg/kg, daily for 5 days, i.p.). As shown in FIG.9, the tumor suppression was greater for the nab-17-AAG plus Abraxane™combination therapy than for either single therapy group.

Although the foregoing invention has been described in some detail byway of illustration and example for purposes of clarity ofunderstanding, it is apparent to those skilled in the art that certainminor changes and modifications will be practiced. Therefore, thedescription and examples should not be construed as limiting the scopeof the invention.

All references, including publications, patent applications, andpatents, cited herein are hereby incorporated by reference to the sameextent as if each reference were individually and specifically indicatedto be incorporated by reference and were set forth in its entiretyherein.

Preferred embodiments of this invention are described herein, includingthe best mode known to the inventors for carrying out the invention.Variations of those preferred embodiments may become apparent to thoseof ordinary skill in the art upon reading the foregoing description. Theinventors expect skilled artisans to employ such variations asappropriate, and the inventors intend for the invention to be practicedotherwise than as specifically described herein. Accordingly, thisinvention includes all modifications and equivalents of the subjectmatter recited in the claims appended hereto as permitted by applicablelaw. Moreover, any combination of the above-described elements in allpossible variations thereof is encompassed by the invention unlessotherwise indicated herein or otherwise clearly contradicted by context.

1. A method of treating a proliferative disease in an individual,comprising administering to the individual an effective amount of acomposition comprising nanoparticles comprising 17-AAG and a carrierprotein.
 2. The method according to claim 1, wherein the proliferativedisease is cancer.
 3. The method according to claim 2, wherein thecancer is lung cancer.
 4. The method according to claim 1, wherein the17-AAG is administered intravenously.
 5. The method according to claim1, wherein the nanoparticles comprise 17-AAG coated with the carrierprotein.
 6. The method according to claim 1, wherein the carrier proteinis albumin.
 7. The method according to claim 6, wherein the albumin ishuman serum albumin.
 8. The method according to claim 1, wherein theaverage diameter of the nanoparticles in the composition is no greaterthan about 200 nm.
 9. The method according to claim 1, wherein theweight ratio of the carrier protein to 17-AAG is less than about 18:1.10. The method according to claim 5, wherein the carrier protein isalbumin.
 11. The method according to claim 10, wherein the albumin ishuman serum albumin.
 12. The method according to claim 10, wherein theaverage diameter of the nanoparticles in the composition is no greaterthan about 200 nm.
 13. The method according to claim 12, wherein theweight ratio of the carrier protein to 17-AAG is less than about 18:1.14. The method according to claim 11, wherein the average diameter ofthe nanoparticles in the composition is no greater than about 200 nm.15. The method according to claim 14, wherein the weight ratio of thecarrier protein to 17-AAG is less than about 18:1.